Molbio Syll

SYLLABUS OF

M.Sc., MOLECULAR BIOLOGY

FROM 2006 - 2007 ONWARDS

SEMESTER – I THEORY: Paper - I Paper – II Paper – III Paper - IV

: : : :

PRACTICALS Practical- I : Practical - II :

MLB 101 MLB 102 MLB 103 MLB 104

: : : :

Biochemistry – I Molecular Physiology Molecular Cell Biology Microbiology

: :

MLB 105 MLB 106

Biochemistry – I and Molecular Physiology Molecular Cell Biology and Microbiology

SEMESTER – II THEORY Paper - I Paper - II Paper - III Paper - IV

: : : :

PRACTICALS Practical- I : Practical - II :

MLB 201 MLB 202 MLB 203 MLB 204

MLB 205 MLB 206

: : : :

Nucleic Acids Genetic Engineering Genetics and Cytogenetics Biostatistics, Computer Applications and Bioinformatics

: :

Nucleic Acids & Genetic Engineering Genetics and Cytogenetics and Biostatistics, Computer Applications and Bioinformatics

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SEMESTER – I Scheme of Study and Examination Code of the Paper

(Hrs / week) Paper

MLB101 MLB102 MLB103 MLB104 MLB105

I II III IV Pract .I

MLB106

Pract .II

Subject Biochemistry – I Molecular Physiology Molecular Cell Biology Microbiology Biochemistry – I & Molecular Physiology Molecular Cell Biology & Microbiology

Theory

Practical

4 4 4 4 --

----8

Total No. of hrs/ Semester 52 52 52 52 104

--

8

104

Duration of examination (hrs) 3 3 3 3 4

Max. Marks of examination 80 80 80 80 40

4

40

Continuous evaluation

Total Marks

20* 20* 20* 20*

100 100 100 100

10**

50

10**

50

TOTAL MARKS

500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical Record + 5 marks for visit to Industries/Laboratories

3

SEMESTER – II Scheme of Study and Examination

Code of the Paper

(Hrs / week) Paper

MLB201 MLB202 MLB203 MLB204

I II III IV

MLB205

Pract .I

MLB206

Pract .II

Subject Nucleic Acids Genetic Engineering Genetics and Cytogenetics Biostatistics, Computer Applications & Bioinformatics Nucleic Acids & Genetic Engineering Genetics & Cytogenetics and Biostatistics, Computer Applications & Boinformatics

Theory

Practical

4 4 4 4

-----

Total No. of hrs/ Semester 52 52 52 52

Duration of examination (hrs) 3 3 3 3

Max. Marks of examination 80 80 80 80

--

8

104

4

40

--

8

104

4

40

Continuous evaluation

Total Marks

20* 20* 20* 20*

100 100 100 100

10**

50

10**

50

TOTAL MARKS

500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical Record + 5 marks for visit to Industries/Laboratories

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QUESTION PAPER FORMAT BANGALORE UNIVERSITY M.Sc., (I & II Semester) Examination MOLECULAR BIOLOGY Paper: Time: 03 Hours

Max. Marks: 80

Instructions: 1. Answer all the parts 2. Draw diagrams wherever necessary A. Write short notes on the following: 1-5

(5x2=10)

B. Write critical notes on the following: 6-10

(5x8=40)

C. Answer any two of the following 11-14

(2x15=30)

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SEMESTER – I MLB – 101 BIOCHEMISTRY – I 52 hrs Unit - I Chemical bonding: Covalent bonds, ionic bonds, hydrogen bonds, co-ordinate bonds; Electrostatic attractive forces, van der Waals forces, hydrophobic forces; Involvement of molecular orbitals in chemical bond formation, sigma and pi bonds; Bond strength, bond energy and bond radius; Formation of chemical bonds with carbon and other elements. Geometry of carbon compounds, tetrahedral structure, conformation and configuration, optical activity, asymmetric centers or chiral centers, stereoisomers, cis-trans configuration. 05 hrs Unit – II pH: Weak acid and weak base; Dissociation constant, ionization of water and equilibrium constant, properties of water, hydrogen ion concentration, Henderson-Hasselbalch equation. Buffers: Concept, importance and preparation. 04 hrs Unit - III Bio-organic reactions : Acid–base , Covalent and Metal ion catalysis; Concept of nucleophiles and electrophiles, Nucleophilic substitution reactions and their importance. 02 hrs Unit - IV Chemistry of Biomolecules: Carbohydrates: Classification, chemistry of mono, oligo and polysaccharides. Functions of homo and hetero polysaccharides. Lipids: Classification, brief outline of the chemistry of fatty acids, phospholipids, sphingolipids. Amino acids: Classification, properties, protein and non-protein amino acids, essential and nonessential amino acids. Modified amino acids and their functions. 06 hrs Unit - V Proteins: Structural Organization and Classification

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Concept of peptide bond; steric interference, Ramachandran plot and its importance. Primary, secondary, tertiary and quaternary structure; Secondary structural forms such as alpha-helix, beta sheet conformation, sequence driven conformation, helix turn helix, beta turn beta and combination of them as structural motifs, structure prediction; Concept of motif and domain; Forces for tertiary folding and its stabilization; Protein-protein interaction, denaturation and renaturation; Determination of isoelectric point; Estimation of protein purity. Protein sequencing methods; Concept of protein microarray methods. specific and molecular activity of proteins. Role of chaperones and isomerases in 3-D folding of proteins. 10 hrs Unit - VI Enzymes: Chemistry and structure: Basic organization 3-D structure of simple enzyme and multimeric enzyme and multienzyme complex; Structural features such as substrate binding site, catalytic site, allosteric site; Mechanism of enzyme activation, induced conformational changes. Cofactors and Activators: Characteristic features of cofactors, coenzymes and their role in chemical reactions, concept of apozymes, prosthetic groups and holozyme. 07 hrs Unit - VII Enzyme Kinetics: Rate of reaction, kinetic order rate equations, turn over, Kcat, Vmax, Km, Michaelis-Menten equation, Lineweaver-Burk plot, activation energy, binding energy, transition states, equilibrium, kinetics of enzyme inhibitors. Mechanism of Enzymatic Catalysis: Principles and concepts of specificity, binding, entropy reduction, desolation, transitional state and induced fit processes; Acid-base catalysis, covalent catalysis, metal catalysis; Single substrate reaction and double substrate reaction. Factors: pH, temperature, concentration of substrates. Pre-pro enzymes and Regulation of Enzyme activity: Concept of feed back inhibition and allosteric regulation. 08 hrs

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Unit – VIII Biochemical mechanisms: Introduction, mechanistic role of the following in living systems :- thiamine pyrophosphate (TPP) in decarboxylation of alpha keto acids and in the formation of alpha ketols; Pyridoxal phosphate (PLP) in transamination, decarboxylation, dealdolisation and elimination reactions of amino acids; Lipoic acid in the transfer of acyl group, co-enzyme A (coASH) in the transfer of acyl group, biotin in the carboxylation reaction; Tetrahydropholic acid in one carbon transfer reaction; Vitamin B12 co-enzymes in molecular rearrangement reactions and in the synthesis of methionine and methane; Nicotine amide and flavin coenzymes in biological redox reaction. Biochemical Techniques: Principle and application of Spectrophotometry, X-Ray diffraction, NMR, Centrifugation, Chromatograpy and Electrophoretic techniques. 10 hrs

Reading References: 1. Boyer, R.F. [Ed.] (1986) Modern Experimental Biochemistry; Addison Wesley 2. Buchanan, B.B., Wilhelm Gurssem & Jones R.L. (2000), Biochemistry & Molecular Biology of plants. American Society of Plant Physiologists, Rock Ville, USA, Maryland. 3. Colowick, S.P. et al., [Eds.] (1987) Methods in Enzymology; Vol. 152, Academic press. 4. Conn, E.E and Stumpf, P.K, G.Bruencing and R.G. Dol (1995). Outlines of Biochemistry. John Wiley, Singapore. 5. Conn, E.E and Stumpf, P.K. (1976), Outlines of Biochemistry, John Wiley and sons Inc, New York 6. Dey, P.M. and J.B.Harborne (1997), Plant Biochemistry: Academic press, Inc.San Dugo, California V 7. Engel, P.C. (1981), Enzyme Kinetics; The steady state approach Champman and Hall 8. Hans – Walter Heldt, (2005), Plant Biochemistry, 3rd edition, Academic Press, An Imprint of Elsevier 9. Irwin H.Segal (1976), Enzyme Kinetics ; Interscience – Wiley 10. Keithwilson and John Walker, (Ed) (2005). Principles and techniques of Biochemistry and Molecular Biology Cambridge University Press. 11. Mathews and Van Holde (1995), Biochemistry 2nd Edition, Benjamin/Cummings publishing company Inc. 12. Nelson, D.L. and Cox, M.M. (2004), Principles of Biochemistry, CBS publishers and Distributors. New Delhi. 13. Roberts , D.V. (1977), Enzyme Kinetics ; Cambridge University Press. 14. Stryer, L.(1995), Biochemistry (4th Edition) W.H.Freeman and company, New York 15. Thomas M. Devlin, (2006) Text Book of Biochemistry with clinical correlations, 6th edition, Wiley – Liss Publication. 16. Voet , D and Voet, J.G. (2004), Biochemistry, 2nd edition J.Wiley and sons 17. Wilson, K and J. Walker (1995), Practical Biochemistry; Principles and Techniques; Cambridge University Press.

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18. Zubey G.L. (1998). Biochemistry, Wn. C. Brown publisher, Sydney 19. Zubey, G.L. Parson, W.W. and Vance, D.E. (1994), Principles of Biochemistry Wn. C. Brown publishers. Oxford.

MLB – 102 MOLECULAR PHYSIOLOGY 52 hrs Unit – I Membrane Transport: Overview, molecules across biomembrane by passive diffusion, protein mediated transport, uniport transport, GLUT1 uniport transport, Human GLUT proteins. Pumps: Different classes, structure and functional properties; ATP-driven ion pumps; Ca2+ ATPase pump, Calmodulin mediated pump, Na+/K+ ATPase pump, V-class H+ ATPase pump. Carriers: Glucose and amino acid transport proteins ABC transporter: Bacterial permeases, ABC small molecular pumps; Flippare mechanism Ion channels: Nongated ion channels, co-transport by symporters and antiporters, voltage gated ion channels. 08hrs Unit – II Protein sorting: The machinery of protein sorting: Translocation of secretary proteins across the membrane of endoplasmic reticulum, insertion of proteins into the membrane of endoplasmic reticulum, protein modification and folding, export of bacterial proteins. Targeting proteins to mitochondria and chloroplast. Vesicle traffic: Secretory pathway, molecular mechanism of vesicular traffic, early and late stages of secretory pathway. 07 hrs Unit-III Exocytosis: Transport of molecules from the trans Golgi network to the cell surface, and role of annexins in exocytosis, Endocytosis: Receptor for low density lipoproteins, ligands for endocytosis, endosomes for dissociation of receptor – ligand complexes, the endocytic pathway, specialized vesicles to deliver cell components to lysosomes, degradation. Transcytosis : Endocytosed ligands across an epithelial cell layer 06 hrs

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Unit – IV Defense system: Plant pathogen interaction, R-genes and R-gene mediated disease resistance. Plant defense reaction and response. Control of pathogens by genetic engineering. Chaperones and Prions. 06 hrs Unit – V Amino acid Metabolism: Amino acids as neurotransmitters, aromatic amino acid biosynthesis, aspartate derived amino acid biosynthesis, proline metabolism: and its utility in evolving stress tolerance in plants. 05 hrs Unit –VI Lipid Metabolism: Fatty acid biosynthesis, desaturases, synthesis of unusual fatty acids, membrane lipids and their function, synthesis and catabolism of storage lipids. 05 hrs Unit – VII Cellular Receptors and Signal Transduction: Concept of cellular signaling, characteristic signaling molecules, extra cellular and intracellular forms, paracrine, synaptic and endocrine form of signaling, concentration and their effects, cellular status and potential (programmed) in receiving and responding to signals, cellular receptors and their characters. Cell surface receptor proteins ionchannel linked, G-protein linked and enzyme linked; Intracellular receptors - kinds and mechanism: mechanism of binding, activation and signal transduction. Role of Inositol Phosphotidyl, diacyl-glycerol molecules and Calmodulin, cAMP and cGMPs, kinases, phosphatases in signal transduction. 10 hrs Unit – VIII Molecular mechanism of plants and animal adaptation to temperature and osmotic pressure with reference to high and low altitude. 05 hrs Reading References: 1. Buchanan, B.B, Gruissem, , W. and Jones, R.L. (2004). Biochemistry and Molecular Biology of plants. I.K. International Pvt., New Delhi. 2. Conn, E.E., Stumpf., Bruenning, G and Doi, R.H. (1987). Outlines of Biochemsitry. John Wiley and Sons, New York. 3. Flochachka and Sumarea. M. (1989). Molecular Mechanism of Adaptations, Academic Press, New York. 4. Gerald Karp. (1996). Cell and Molecular Biology – Concepts and Experiments. John Wiley and Sons, Inc., New York. 5. Gupta, P.K. (2004). Cell and Molecular Biology. Rastogi Publications, Meerut.

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6. Harvey Lodish, Arnold Berk, Paul Matsudaira, Chris A. Kaiser, Monty Krieger, Matthew P. Scott, S. Lawrence Zipursky and James Darnell. (2003). Molecular Cell Biology, W.H. Freeman and Company, New York. 7. Hopkins, W.G. (1995). Introduction to plant physiology. John Wiley & Sons Inc. New York, USA. 8. Moore, T.C. (1989). Biochemistry and physiology of plant hormones. 2nd edition. Springer-Verlag, New York , USA. 9. Stumpf, P.K. and Conn, E.E. (Eds) (1988). The biochemistry of plants- A Comprehensive treatise. Academic press, New York. 10. Schmidt – Nelson, R. (1987). Annual Physiology, Adaptation and Environment, 5th Edition, Cambridge University Press, London 11. Taiz, L. and Zeiger, E. (2003). Plant Physiology. 3rd edition. Panima Publishing Corporation, New Delhi/Bangalore 12. Taiz, L. and Zeiger, E. (1998). Plant Physiology. 2nd edition. Sinauer Associates, Inc., Publishers, Massachusetts, USA. 13. Wilkins, M.B. (eds.) (1989). Advanced Plant Physiology. Pitman Publishing Ltd., London.

MLB – 103 MOLECULAR CELL BIOLOGY 52 hrs Unit – I Cell wall: Cell wall composition, biosynthesis and assembly. Membrane structure - Fluid mosaic model. Ultra structure and function of membranous organelles: Endoplasmic Reticulum, Golgi Apparatus, Vacuoles, Lysosomes, Chloroplast and Mitochondria 08 hrs Unit - II Cytoskeleton: Microfilaments, Microtubules and Intermediate filaments. Actin and tubulin gene families, dynamics of actin and tubulin, role of microtubules in intracellular movement, mitosis and cytokinesis 06 hrs Unit - III Ribosomes: Structure of prokaryotic and eukaryotic ribosomes. Riboproteins, rRNA , 3-D modular structure. Ribosome biogenesis. Important structural motifs in ribosomes and their functions. 06 hrs Unit - IV Nucleus: Nuclear membrane, nuclear pore complex, transport mechanism of nuclear laminins and their role. Composition of nuclear sap, nucleolar sap and nucleoplasm.,

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Nucleolus: Organization, chemistry, function and biogenesis

05 hrs

Unit – V Molecular organization of Eukaryotic chromosome: Molecular organization of nucleosome, nucleosomal thread, solonoid and coiled coil structures. Molecular features of telomere, kinetochore, euchromatin and heterochromatin; Role of scaffold and matrix proteins (SARS and MARS); Mechanism and factors for chromosomal condensation and relaxation. 08 hrs Unit – VI Chloroplast: Genome size, number, organization of genes; Chloroplast mRNA and tRNA, ribosomes, biogenesis of plastids, role of light and phytochrome in the development of plastids; Import of proteins from cytoplasm. Mitochondria: Genome size, number and organization of genes; Molecular organization of inner and outer membrane, attachment sites and their function; mitochondrial matrix and its chemical composition and physiological functions. Organization of electron transport system in the inner membranes, elementary particles and their organization and functions. Mitochondrial protein synthesis; Transport mechanism of cytosolic proteins into mitochondria, Biogenesis of mitochondria. 08 hrs Unit – VII Cell Cycle: Genetic regulation of cell cycle; Molecular basis of cellular check points. 05 hrs Unit - VIII Apoptosis: Programmed cell death (PCD), signals and causes for inducing apoptosis, regulation of gene activity during apoptosis, role of cysteine-containing-aspartate specific proteases (CASPASes), Apoptotic Protease Activating Factors (APAF) mechanism of apoptosis at biochemical level, importance of apoptosis in development and organogenesis. Programmed cell death (PCD) in plants and its relation to senescence, senescence associated genes (SAGs), hypersensitivity as a cause for apoptosis, Necrosis, tumour necrosis factor, death signals, cell death by activating Caspases 06 hrs Reading references: 1. Alberts. B., Bray, D., Lewis, J., Raff, M., Roberts, K and Watson, J.D. (1994). Molecular Biology of the cell. Garland Publisher Inc., New York 2. Bishop J.A. (1982). Retroviruses and cancer genes. Advances in cancer research.

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3. 4. 5. 6. 7. 8.

Brachet J. (1985). Molecular Cytology . Academic Press New York Celis J E (Eds): Cell Biology : A Laboratory Hand Book . Vol I & II Academic Press. Elliot and Elliot. (2001). Biochemistry and Molecular Biology. Oxford University Press. Gerald Karp. (1996). Cell and Molecular Biology. John Wiley and Sons. Inc J.Daneil H and Lodish D. (1995). Molecular cell Biology. Baltimore Scientific American Book Knudson A.G. (1998). Anti – Oncogenes and Human cancer . Proceedings of the National academy of xciences USA 90:10 0114-10921 9. Lodish,H., Ber, A., Zipuoskry, L.S., Matsudaira, P., Bahimore, D and Damell J. (2001) Molecular Biology W.H Freeman G Co. 47 10. Pollard J.P. and W.C. Earnshaw. (2002). Cell Biology, Sunders 11. Sudberry P. (2002). Human Molecular Cytogenetics. Prentice Hall Publication 12. Wolfe. A. (1995). Chromatin structure and function. Academic press; New York

MLB – 104 MICROBIOLOGY 52 hrs Unit - I Introduction to viruses: Structure, properties, importance and classification. Bacterial viruses: Structural organization, replication and assembly of dsDNA phages: T4 phage, T7 phage and Lambda phage; ssDNA phages: Øx 174 phage and M13 phage; RNA phages: Ms2 phage Plant viruses: Occurrence, distribution, structural organization of capsid and genome organization and replication, disease caused by the viruses and methods to contain the viruses. DNA viruses: Gemini Virus, Cauliflower Mosaic Virus. RNA viruses: Tobacco Mosaic Virus, Potato Virus - X and Y, Gamphrena Virus (Rhabdovirus), Plantain-B Virus. 10 hrs Unit - II Bacteria: Occurrence, distribution, shape, size, pathogenesis of bacteria and control measures. Gram negative and positive bacteria. Ultra structure of E-coli. Basic mechanism of bacterial membrane transport : Passive, facilitated passive transport, Active transport-pumps, carriers; group translocation, translocation of sugars, ions, amino acids, and their regulation. Flagella and Cilia: Structural

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organization of flagella and cilia and molecular mechanism of movements. Fimbriae and sex pili: Structural organization and functions of Fimbriae and Sex pili; Chromosome: Genome size and organization (nucleoids). Cell division: Molecular Mechanism of cell division and its regulation; genes involved in cell division. 08 hrs. Unit - III Recombination in bacteria: a) Conjugation, Discovery, nature of donor strains and compatibility, interrupted mating and temporal mapping, Hfr, F’, heteroduplex analysis, mechanism of chromosome transfer, molecular pathway of recombinations and genemapping. b) Transformation in Gram +ve and Gram –ve bacteria, natural transformation systems, biology of transformation, transformation and gene mapping, chemical mediated and electro tansformations c)Transduction Discovery, generalized and specialized transduction, phage P1 and P22 mediated transduction, mechanism of generalized transduction, abortive transduction, mechanism of specialized transduction, gene mapping, fine structure mapping. 05 hrs Unit - IV Agrobacterium: Morphology, Ti plasmids, and T-DNA, their characteristic features, mechanism of T-DNA transfer into plant cells, Ti- mediated crown gall development, mechanism of plant cancer development. Bacterial plasmids: Characteristic features and classes of plasmids- sex factor F+/- containing plasmids, R-plasmids, Pathogenic plasmids (K-plasmids, Hly and Enteric plasmids), Col-plasmids, Degradative plasmids, Tumour inducing plasmids, Cryptic plasmids, metal resistance plasmids- and their characters. Transmission of Colicin and drug resistant R plasmids. 04 hrs Unit - V Cyanobacteria: Structural features, cell structure, organization of membranes for photosynthesis, nitrogen fixation, genomes and reproductive methods and use of cyanobacteria as fertilizers. Transposable elements: Insertional elements (IS), Tn elements (Transposons); Structural features, numbers, sizes, replication and mechanism of transpositions, Transposon mediated drug resistance. 04 hrs Unit - VI Fungi: Yeast: Detailed account of Saccharomyces cervisiae, cell biology, reproductive biology, genetics of mating types, nuclear and cytoplasmic inheritance. Importance of Saccharomyces and its related members in genetic engineering and commercial uses.

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Mycorhizae and Vam fungi: Composition, structural features and their importance. 05 hrs Unit - VII Microbial metabolism: Bacterial photosynthesis: Phototrophs, .chemotrophs, chemical composition of photosynthetic components. Mechanism of light reaction and dark reaction - Reductive pathway, pentose pathway and pyruvate synthase pathway. Mechanism of chemosynthesis. Bacterial carbohydrate metabolism: Embden-Meyerhoff and Parnas pathway, Entner-Duodorff pathway and Warburg-Dickens pathway, Pentose and Hexose phosphoketolase pathway. Aerobic pathways leading to citrate cycle and oxidative phosphorylation 06 hrs Unit - VIII Genetically Engineered Microbes (GEMS): Gene manipulated microbes, Production of therapeutic agents: Gene engineering and production of - Human interferons, human growth hormones, DNase-I, alginate lyase. Production of humanized and regular antibodies in E.coli and yeast. Anti HIV agents. Use of GEMS for the production of proteins, antibodies and other metabolites of importance. Biofertilizers: Use of microbes and genetically engineered microbe in enriching soil with fertilizers. Nif genes: Nitrogen fixing genes and organization, Genetic engineering of Nitrogen fixing gene clusters in Klebsiella pneumoniae. Regulation of nif genes. Hydrogenase genes: Cloning and expression of Hup+ genes (Hydrogenase gene) and its importance. Genetic engineering of nodulation genes and its importance in agriculture. Cloning of antibiotic genes from pseudomonads: Cloning and expression of antibiotic genes and its importance. 10 hrs Reading References: 1. Atlas R.M. (1998) Microbiology : Fundamental and application (IIeds) Mac millan Publishing company 2. Bruijin et al ., (1998). Bacterial genomes, Chapman and Hill 3. Dale J.W. (1994). Molecular genetics and Bacteria. John Wiley and sons 4. Hayes W. (1970). The genetics of Bacteria and their viruses. The English Book society of Blackwell Scientific Publication, Oxford

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5. 6. 7. 8.

Hunderson et al., (1999). Cellular Microbiology Wiley Lewin B (2002). Genes VIII, Oxford Prescot L.M., Hanley, J.P. and Klein, D.A. (1999). Microbiology, WCB Mc Graw Hill , Con MY Roger L.P., John T., Knowler and Daviol P. Leadr. (1992). The Biochemistry of Nucleic acids, 11th edition. Chapmann and Hall 9. Samuel Singer (2001). Experiments in Applied Microbiology, Academic Press New York. 10. Stnely R. Maloy, John E. Cronan, Jr., David Freifelour (1994). Microbial genetics. Jones and Barlett Pub. Bosten. 11. Sullia S.B. and S.Shantharam (1998). General Microbiology , Oxford IBH Publishing Con, New Delhi.

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FIRST SEMESTER PRACTICALS MLB – 105 BIOCHEMISTRY – I AND MOLECULAR PHYSIOLOGY PART – A Preparation of Buffers Protein isolation (crude) from plant, animal and microbial sources Estimation of Protein – Colorimetric method Estimation of amino acids – Ninhydrin method Determination of Specific activity of Enzyme- Invertase Determination of Km using Line weaver – Burk plot PART – B 7. Extraction of lipids - Plant and animal sources 8. Salicylic acid chromatography of lipid extracts 9. TLC of lipids and identification of different lipids 10. Separation of amino acids by paper chromatography and TLC 1. 2. 3. 4. 5. 6.

MLB – 106 MOLECULAR CELL BIOLOGY AND MICROBIOLOGY PART – A 1. Preparation of meiotic chromosomes using Haemotoxylin/Feulgen stain – Poecilocera picta 2. 3. 4. 5. 6.

Vital staining of mitochondria Isolation of mitochondrial DNA Isolation of chloroplast DNA Preparation of salivary gland chromosomes – Drosophila melanogaster Vital Staining – Animal and Plant, Dye exclusion technique PART – B 7. Growth curve of E.coli 8. Isolation of bacterial plasmids

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9. Isolation of cyanobacteria 10. Bacterial respiration 11. Estimation of oxygen during photosynthesis 12. Plasmid transformation

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SECOND SEMESTER MLB – 201 NUCLEIC ACIDS 52 hrs Unit - I Chemistry and properties of Nucleic Acids: Discovery of genetic material, chemistry and structural features of nucleotides. Quantitative relationship between bases, ionic state, sugar puckering, base configuration with respect to the sugar. Comparative properties of DNA and RNA with respect to size, quantity, distribution, turnover, Watson-Crick base paring, Non-Watson–Crick base pairing by hydrogen bonds. Procedures and protocol for DNA melting and annealing by DNA:DNA hybridization and DNA:RNA hybridization, kinetics of melting and reannealing. Cot curves and Cot ½ values and rot and rot ½ curves, kinetic complexity and chemical complexity. Importance of kinetic class of DNA. 08 hrs Unit - II Structural forms of DNA: dsDNA: (ds= double stranded) Structural features of Watson-Crick B-DNA model, the basis for A=T and G=C base pairing by hydrogen bonds, nature of sugar phosphate backbone, projection of bases, hydrophobic interaction between stacking of base pairs, tortional twist, thermodynamic compatibility. Structural features of Z-DNA and A-DNA and important differences between B, Z and A forms and their importance. ssDNA: (ss = single strand) the nature and properties of ssDNA and its structural form. tsDNA: (ts = triple strand) Structural features of tsDNA, sequences responsible for forms, inter and intra-strand triple helices and disease due to tsDNA formation. qsDNA: (qs = quadruplex strand): Characteristic features of four stranded DNA, basis for four stranded forms, the structure and the role of telomeric DNA. Supercoiled DNA: Structural features of negative and positive supercoiling. Role of topoisomerases in maintaining the supercoiling. Other forms of DNAs: Structural distinguishing features of Cruciform DNA, Flexible DNA, Curved DNA and their importance. 08 hrs Unit - III Replication of DNA:

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Replication of single stranded DNA: Mechanism of ssDNA replication of Ø x174 and M13 phage; structural features of origin, primosomal assembly and primosomal complex, factors and DNA-polymerases , replication mechanism, and the enzymes involved. Replication of Prokaryotic DNA: Replicons, number of replicons per genome. Structural feature of E.coli genomic Origins (ORI) and Termination regions (TER). Factors and characteristic features of different DNA-polymerases involved in replication apparatus (replisome); subunits and their functions, primosomal assembly, primer formation, assembly of subunits into functional DNApolymerase-III. Mechanism of replication: Replication of leading strand and replication of lagging strand, fidelity, rate, and regulation of replication in bacteria. Repair of replication error. 09 hrs Unit - IV Replication of Eukaryotic DNA: Organization of chromosomal replicons, number and structural features of eukaryotic origins with examples from yeast plasmids and yeast chromosomes. Characteristic features of DNA polymerases – alpha and delta, their chemical composition and associated subunits. DNA-Pol associated factors : RF-A, RF-C, PCNA, MF-1 and their role. Mechanism of replication : Replication of telomeric DNA – telomerase, components and the mechanism. 07 hrs Unit – V Genetic Code: The language of information transfer, genetic and biochemical analysis of genetic code, Features of genetic code and evolution of genetic code DNA damage and Repair: Causes for DNA damage. Kinds of errors in DNA: natural, chemical and radiation, their effects as heritable mutations, frequency of mutations, kinds of mutations and their importance in evolution. Repair mechanism of DNA damage, genes and the factors involved in repair mechanism. 07 hrs

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Unit - VII Chemistry and Structural forms of RNA: Characteristic features and classification of RNA. Genetic RNA: A brief review of RNA viruses, structure and replication of RNA genome: Picorna virus, Rabies virus and Retrovirus (HIV). Viroidal and virusoidal RNA: Structure and characteristic features of viroids and their RNA. Concept of ribozyme and mechanism of ribozyme action. Characteristic features of few satellite viral RNAs. Ribosomal RNAs: Structural features of prokaryotic and eukaryotic rRNA systems, size, kinds of rRNA and base modifications. Precursors and processing, assembly and organization of the rRNA into ribosomes, important structural motifs and their functions 06 hrs Unit - VIII tRNA: Structure and nature of prokaryotic and eukaryotic tRNAs, base modifications, organization of tRNA into 3-D structure, different motifs and their functionl features. Aminoacylations, aminoacyl-tRNA synthetases and their features, concept of second codon, mechanism of aminoacylation of tRNA. Anticodons and operation of Wobble mechanism and its importance. Precursor tRNAs and processing of tRNAs. snRNAs, scRNAs and ncRNAs: Structure, kinds, genes, their associate proteins and the function of snRNAs, scRNAs and snoRNAs in precursor RNA processing miRNAs, siRNAs and tmRNAs mRNAs: Structural features of prokaryotic and eukaryotic mRNA. Concept of introns and exons, exons as protein motifs and their origin and evolution. 07 hrs Reading References: 1. David. A. Micklos, Greg.A. Freyer and David A. Crotty, (2003). DNA Science A First Course, 2nd edition, Cold Sprind Harbor Laboratory Press, New York. 2. Flint. S.J, L.W. Enquist, R.M. Krug, V.R. Racaniello and A.M. Skalka, (2000) Principles of Virology, ASM Press, Washington D.C 3. Gerald Karp (1996). Cell and Molecular Biology – Concepts and Experiments. John Wiley and Sons, Inc., New York. 4. Gupta, P.K. (2004). Cell and Molecular Biology. Rastogi Publications, Meerut. 5. G.M. Malacinski and D. Friefelder (2005). Essentials of Molecular Biology vol – I, Jones and Bartlet Publishers. 6. Griffiths AJF, H.J. Muller., D.T. Suzuki, R.C. Lewontin and W.M. Gelbart (2000). An introduction to genetic analysis. W.H. Freeman , New York 7. Harvey Lodish, Arnold Berk, Paul Matsudaira, Chris A. Kaiser, Monty Krieger, Matthew P. Scott, S. Lawrence Zipursky and James Darnell. (2003). Molecular Cell Biology, W.H. Freeman and Company, New York.

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8. Lewin B. (2003). Genes VIII Oxford University Press. Oxford 9. Miglani G.S. (2002). Advanced Genetics, Narosa Publishing House , New Delhi. 10. Watson, Baker, Bell, Gann, Levine and Losick. (2006). Molecular Biology of the Gene, 5th edition, Pearson Education. 11. Watson, J.D. T.A.Baker, S.P. Bell, A.Lann. M.Levine and R.Losick. (2004). Molecular Biology of genes, V edition, Perason Education RH Ltd., India

MLB – 202 GENETIC ENGINEERING 52 hrs Unit – I Introduction: Phases of genetic studies, historical account, definition and objectives 02hrs Unit – II Molecular tools, restriction and modifying enzymes, other nucleases, polymerases, ligase, kinases and phosphatases. Isolation and purification of DNA and RNA. 06 hrs Unit – III Cloning Vectors: Plasmids, phages, cosmids, artificial chromosomes and expression vectors. Cloning hosts: E.coli, Saccharomyces, plant and animals cells. Cloning protocols: General transformation, electroporation, pofectamine, gene-gun method. 06 hrs

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Unit – IV Gene isolation: Methods of construction of genomic and cDNA libraries, direct cDNA cloning, positional cloning. RFLP mapping, chromosome walking and jumping 06 hrs Unit – V Screening and characterization of clones: Molecular probes and methods of labelling, principles of hybridization and hybridization based techniques (colony plaque, Southern, Northern and in situ hybridizations). Expression based screening, 07 hrs Unit – VI DNA sequencing: Maxam and Gilbert’s method, Sanger’s method, Automated DNA sequencing, Capillary gel electrophoresis method and DNA sequence analysis. 05 hrs Unit – VII Polymerase chain reaction: Mechanism, methods-RT-PCR, RAPD, AFLP, ISSR, real time PCR and its application 08 hrs Unit – VIII DNA Engineering techniques: Gel electrophoresis of nucleic acids (denaturing and native), gel electrophoresis of proteins (SDS PAGE), pulse-field gel electrophoresis of DNA, oligonucleotide synthesis, Microarray technology, RNAi technology, blotting of macromolecules, Promoter characterization : Promoter analysis through reporter genes, electrophoretic mobility shift assay, DNA foot – printing, DNA fingerprinting, Mutagenesis: Site directed mutagenesis, Transposon mutagenesis, Construction of knock-out mutants, Gene transfer techniques (Physical and vector mediated methods), Transfection of cells: Principles and methods, Germ line transformation in Drosophila and transgenic mice : Stratagies and methods, In vitro translation 12 hrs

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Reading Reference 1. Brown,T.A. (1995). Gene Cloning: An introduction . Chapman and Hall, London 2. Glick,B.R. and Pastenak, J.J. (1994). Molecular Biotechnology: Principles and applications of recombinant DNA. ASM Press, Washington D.C. 3. Kreuzer, H. and A. Massey. (2001). Recombinant DNA and Biotechnology. ASM press, Washington DC 4. Lodish, Berk, Zipursky, Matsudira, Baltimore and Darnell. (2005) Molecular cell Biology, W.H.Freeman and Company 5. Mathew R Walker, Ralph Rapley. (1997) Route maps in Gene Technology, Blackwell Publishing. 6. Micklos, D A, Freyer GA and Crotty D A (2003) DNA Science, Second edition, Cold Spring Harbour Laboratory Press, New York. 7. Primrose, S.B., R.M. Ywyman and R.W. Old. (2006). Principles of Gene manipulation and tenomics Seventh edition, Blackwell Science,U.K. 8. Watson, Baker, Bell, Gann, Levire and Losick, (2005), Molecular Biology of the Gene, 5th edition, Pearson Education Publication.

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MLB – 203 GENETICS AND CYTOGENETICS 52 hrs Unit - I Chromosomal organization: Chromosome number, size, morphology, chromatid, centromere, telomere, secondary constriction, chromomere, heterochromatin. Special chromosomes: Lampbrush chromosomes, Polytene chromosomes and Accessory chromosomes. 08 hrs Unit -II Molecular mechanism of cell division: Molecular organization of centrosome and spindle, dynamic instability of microtubules, kinetochore and microtubule interaction, synaptic pairing, chromosome movements during metaphase and anaphase, spindle without chromosomes and cytokinesis. 06 hrs Unit - III Molecular basis of sex determination: Molecular basis of sex determination and dosage compensation in Caenorhabditis elegans, Drosophila and human. 04 hrs Unit -IV Somatic cell genetics: Tissue culture and cell fusion, human and rodent cell hybrids and alignment of gene to chromosomes. Construction of fine scale map. 05 hrs Unit -V Imprinting of genes, chromosomes and genomes: Definition, exception to the principle of equivalence of reciprocal hybrids, pronuclear transplantation experiments in mouse, uniparental chromosomal disomies in mouse, human triploids, sex determination in Coccids, X-chromosome inactivation in marsupial females and molecular mechanisms. 07 hrs Unit - VI Mutations: Key concepts. Forward mutations: At DNA level – Transition and Transversion; At protein level – Silent, missense, nonsense, frameshift mutations, Reverse mutations: Exact reversion, equivalent reversion. Intragenic suppressors: Frameshift of opposite sign and second site within a gene, second site missense mutation. Extragenic suppression: Nonsense suppression, missense suppression, Frameshift suppression, Physiological suppression. Lethal mutation, loss and gain of functional mutations, amorphic, hypomorphic and isoallelic mutations. Chromosome based mutations

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Chemical mutagens: Base analogues: Nitrous acid, hydroxylamine, hydrazine, alkylating agents. Detection of mutations: Bacteria – replica plating technique, Ames test; Drosophila – Sex – linked recessive lethals, autosomal recessive lethals, dominant lethal test; Small mammals – Micronucleus test, dominant lethal assay and host mediated assay. 10 hrs Unit - VII Biochemical genetics of Neurospora: Tetrad analysis and linkage detection in Neurospora, two point and three point crosses, chromatid and chiasma interference. Induction and detection of biochemical mutations in fungi, mitotic recombination in Neurospora crassa and Aspergillus nidulans. Transposable elements in fungi and gene conversion. Biochemical genetics in Algae: Fine structure of an algal cell, algal chromosomes and ploidy. Chlamydomonas – unordered tetrad analysis – recombination and mapping. Nucleocytoplasmic interactions and gene expression in Acetabularia and plastid inheritance. Biochemical genetics of Protozoa: Ultrastructure of a protozoan cell. Reproduction and recombination in Plasmodium. Expression of virulence factors in Plasmodium. 07 hrs Unit – VIII Molecular population genetics: Patterns of change in nucleotide and amino acid sequences, molecular evolution, molecular clock and emergence of Non-Darwinism Molecular phylogenetics: Construction of phylogenetic tree. Phylogenetic inference : Distance methods, parsimony methods and maximum likelihood method. Phylogenetic considerations based on nucleotide and amino acid sequences. 05 hrs

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Reading references:

1. Alberts, B, Johnson, J Lewis, M.Raff, K Roberts and P.Watter. (2002). Molecular Biology of the cell IV eds Garland Science, New York 2. Beatty,, B.S. Mai and J. Squire (2002). FISH. Oxford Univ. Press, Oxford 3. Chatterjee, R.N. (1998) Mechanisms and Evolutionary origins of gene dosage compensation. In Genome analysis in Eukaryotes. Eds. R.N. Chatterjee, and L.Sanchez. Narosa Publishing House, New Delhi 4. Dobzhansky Th., F.J. Ayala,, G.L. Stebbins and J.M. Balentine, (1976). Evolution. Surjeet Publication, Delhi 5. Futuyma D.J. (1986). Evolutionary Biology, Sinauer Associates, INC, Sunderland 6. Hollander A (Editor) (1971-76) Chemical mutagens: Principles and methods of their detection. Vols.1-3, Plenum press New York 7. Lodish, Berk, Matsudaira, Kaiser, Krieger, Scott, Zipursky and Darnell (2005) Molecular Cell Biology, 5th Editon, W.H. Freeman and Company, NY 8. Macgregor, H.C. (1993). An introduction to Animal Cytogenetics, Chapmann and Hall, London 9. Singh, B.D. (2003). Genetics. Kalyani Publishers, New Delhi. 10. Smith, J M (1998). Evolutionary Genetics, Oxford Univ. Press, Oxford 11. Snustad D P, M J Simmons and J P Jenkins. (1997). Principles of Genetics. John Wiley and Sons, INC 12. Verma R.S. (Editor) (1988) Heterochromatin: Molecular and Structural aspects. Cambridge University Press, Cambridge.

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MLB - 204 BIOSTATISTICS, COMPUTER APPLICATIONS AND BIOINFORMATICS 52 hrs BIOSTATISTICS: Unit – I Population and sample, variable, constant and parameters, types of data, summarization of data through frequency distribution, histograms, bar diagrams, and cumulative frequency curve. Measures of central tendency: Mean, median and mode. Measures of dispertion: Variance, standard deviation, co-efficient of variation, quartiles, percentiles and their graphical determination. SkewnessPearson’s and Bowley’s measures. 05 hrs. Unit-II Bivariate Data: Scatter plot, correlation, co-efficient, properties, fitting linear regression, regression co-efficient and interpretation. Probablity: Rules, conditional probability, independence of events, Bayes formula. Sampling: Sampling methods, standard errors of sample means. 05 hrs Unit-III Hypothesis Testing: Basic concepts, large sample tests for proportion, equality of two proportions and means of normal population, confidence intervals for mean and proportion, student’s – test, chi-square test of independent of attributes. ANOVA for one way and two way classification. Dunnett’s and Ducan multiple comparision test. 06 hrs COMPUTER APPLICATIONS: Unit-IV Computer fundamentals, computer organization, computer hardware and computer software, programming languages, operating system, input and output devices, computer memory. 05 hrs Unit-V Word processing, spread sheet calculations and databases. applications. 03 hrs

An overview of MS office.

Computer networks, internet and its

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BIOINFORMATICS: Unit - VI An overview: Applications of Bioinformatics, Data available from various sources, data integration and analysis. Tools of bioinformatics: Web based softwares, commercial softwares Molecular Biology and Bioinformatics: Systems approach to the central dogma of molecular biology. Definitions of various terms and their meaning and relation to bioinformatics. Molecules of Information: A brief account of nucleic acids, DNA, RNA, synthesis if nucleic acids, genes, organization of the same in chromosomes, cloning methods, PCR and DNA sequencing and concept how and in what form the information is encrypted. 10 hrs Unit - VII Proteins: Introduction to protein structure : Secondary and tertiary structure prediction, protein 3D – structure analysis, protein modeling. Principles of homology and comparative modeling Biological database: Types of database, network and database, biological significance of database 06 hrs

Unit – VIII Molecular mapping: DNA sequencing, gene mapping, application of mapping, DNA microarrays, design and data analysis, algorithm in assembling sequence fragments. QTL mapping, candidate gene mapping, physical mapping Proteomics: Proteomic analysis, and tools used, metabolic pathways, genetic network, network properties, simulation of pathway. Phylogenetic analysis: Concept of trees, phylogenetic trees, distance matrix methods, character based methods, evaluation methods. Working with phylogenetic trees. Prediction methods: Use of patterns, methods and tools to predict genes. Prediction of protein structure. Introduction to human genome project. Pharmacogenomics and drug designing. 12 hrs Reading references: 1. Attwood, T.K. and Parry-Smith, D.J. (1999). Introduction to Bioinformatics, Pearson Education 2. Bergeron. (2005). Bioinformatics computing. Pearson Education.

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3. Campbel, (2004). Discovering Genomics, Proteomics and Bioinformatics, Pearson Education. 4. Daniel. (2002). Biostatistics, John Wiley, ISE. 5. Higgins, D and W.Taylor (2000). Bioinformatics Sequence, Structure and databanks , Oxford University Press, Oxford 6. Leibler, D.C. (2002). Introduction to Proteomics, Tools for the new biology, Humana Press, New Jersey 7. Leon and Leon (1999). Information Technology, Leon and leon Publications, Chennai. 8. Lesk, A.M. (2002). Introduction to Bioinformatics, Oxford University Press, Oxford 9. Misener, S and S.A. Krawetz, (2000). Bioinformatics: Methods and protocols , Humana Press, New Jersey 10. Mount, D.W. (2001). Bioinformatics: Sequence and genome analysis. Cold Spring Harbour LaboratoryPress, Cold Spring Harbour New York 11. Primrose S.B., Twyman, R.M. (2006). Principles of Gene Manipulation and Genomics, 7th edition, Blackwell Publishing. 12. Rashidi, H.H. and L.K. Buchler (2000). Bioinformatics Basics: Applications in Biological Science and Medicine, CRC Press, New York 13. Shortliffe, E.H. and L.E.Perreault (Eds.) (2001). Medical informatics: Computer applications in health care and Biomedicine. Springer-Verlag, N.Y. 14. Zar, H.A. (1999). Biostatistical analysis , Person Education

SECOND SEMESTER PRACTICALS

MLB – 205 NUCLEIC ACIDS AND GENETIC ENGINEERING 1. 2. 3. 4. 5.

PART-A Extraction of genomic DNA from plants, animals and microorganisms Restriction digestion of DNA Agarose gel electrophoresis of restriction fragment PCR amplification of DNA Extraction and separation of RNA PART-B

6. Preparation of cDNA library

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7. Cloning and subcloning 8. Screening of cDNA library 9. Blotting techniques: Western blotting and Southern blotting 10. DNA fingerprinting : RAPD assay 11. Restriction site mapping 12. DNA sequencing.

MLB – 206 GENETICS AND CYTOGENETICS AND BIOSTATISTICS, COMPUTER APPLICATIONS AND BIOINFORMATICS PART-A 1. 2. 3. 4. 5. 6.

Culture of Drosophila . Studies on inversion polymorphism in Drosophila Bacterial culture and preparation of competent cells Identification of mutants in Drosophila Studies on phylogenetic trees G and C banding techniques

PART-B 7. Statistical analysis such as descriptive statistics, regression, t-test, Analysis of Variance, multiple regression using statistical software such as SPSS, MINITAB 8. Tests of significance based on Normal, t, Chi-square and F-distributions 9. Correlation and regression

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10. Spread sheet and statistical calculations using MS, EXCEL 11. Internet, E-mail, browsing and searching 12. Practical application of BLAST 13. RNA Folding 14. Identification of genes in genomes 15. Multiple alignment of sequencing

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SYLLABUS FOR III AND IV SEMESTER M.Sc., MOLECULAR BIOLOGY SCHEME OF STUDY AND EXAMINATION SEMESTER III (Hrs / week)

Paper Code

Paper

MLB301

I

Biochemistry – II

MLB302

II

Molecular Biology – I

MLB303 MLB304 MLB305 MLB306

Practical

4 4

--

Title of the Paper

Microbial Biotechnology Molecular Biology of IV Development Pract.I Of paper I & II Pract.II Of paper III & IV III

Theory

Total No. of Hrs/ Semester

Examination

52

Duration (Hrs) 3

Max. Marks 80

Continuous Evaluation 20*

Total Marks 100

--

52

3

80

20*

100

--

52

3

80

20*

100

4

--

52

3

80

20*

100

---

8 8

104 104

4 4

40 40

4

10** 10** Total Marks

50 50 500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical record + 5 marks for visit to Industries/Laboratories

33

SCHEME OF STUDY AND EXAMINATION SEMESTER IV (Hrs / week)

Paper Code

Paper

MLB401 MLB402 MLB403

I II III

MLB404 MLB405 MLB406

Theory

Practical

Total No. of Hrs/ Semester

4 4 4

----

4 ---

Title of the Paper

Immunology Molecular Biology – II Genomics and Proteomics Plant & Animal IV Biotechnology Pract.I Of paper I & II Pract.II Of paper III & IV

Examination

52 52 52

Duration (Hrs) 3 3 3

Max. Marks 80 80 80

Continuous Evaluation 20* 20* 20*

Total Marks 100 100 100

--

52

3

80

20*

100

8 8

104 104

4 4

40 40

10** 10** Total Marks

50 50 500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical record + 5 marks for visit to Industries/Laboratories

1. TOUR: Tour marks should be considered under Continuous Evaluation. a) There will be a Compulsory Industries / Laboratories visit in all the semesters as per relevance to the subject. b) There will be a compulsory Tour 5 to 6 days to any of the industries outside Bangalore in IV Semester.

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THIRD SEMESTER MLB 301: BIOCHEMISTRY - II 52 hrs Unit I: Photosynthesis : Introduction, photosynthetic apparatus – structure, organization, LHC molecules and antenna molecules. Light reaction - cyclic and non-cyclic photophosphorylation. Biochemical reactions –C 4 and C3 pathways, CAM Pathway, Energetics of Calvin pathway. Bacterial photosynthesis. Photorespiration - organells involved, biochemical pathway and its significance. 8hrs Unit II: Carbohydrate metabolism – Glycogen – degradation and biosynthesis. Glycolytic pathway, regulation of glycolysis, gluconeogenesis, hexose interconvertions. HMP pathway. 5hrs Unit III: TCA cycle – pathway and energetics. Alternate pathways – glucuronate pathway and glyoxylate pathway 5hrs Unit IV: Lipids: Definition, Classification, Structure and biological role of Fatty acids, Acyl glycerols, Phospholipids, Glycolipids, Steroids, Prostaglandins, Thromboxanes and Leukotrienes. 6hrs Unit V: Lipid Metabolism: Degradation of triacyl glycerols and phospholipids. Oxidation of saturated, unsaturated fatty acids. Alternate routes of fatty acid degradation. Synthesis of triacylglycerols, phospholipids and biosynthesis and degradation of cholesterol. Metabolism of Prostaglandins and related compounds. 8hrs Unit VI: Thermodynamics – Concept of enthalpy and entropy, free energy and chemical potential. Free energy change, significance of free energy change. Laws of thermodynamics and their applications 4hrs Unit VII:

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Biological oxidation: Basic concept of metabolic energy capture and transfer. High energy compounds – definition, examples. Stages in the production of energy. Biological redox couplers, Redox potentials, Oxidation-reduction reaction free energy changes in electron transfer reactions. 8hrs Unit VIII: Mitochondrial electron transfer system – topology of electron carriers, chemical nature of electron carriers, sequence of electron carriers, isolation of mitochondrial complexes, reconstitution experiments and study of specific inhibitors of ETC. Oxidative phosphorylation – ATP synthesizing system – F0-F1 ATPase. Coupling of electron transfer to ATP synthesis, study of the effects of uncouplers, inhibitors and ionophores, mechanism of oxidative phosphorylation. 8hrs

References: 1. Bob B. Buchanan, Biochemistry and Molecular Biology of Plants (2004), Wilhelm Guissem and Russel L. Jones, I.K. International Pvt. Ltd, New Delhi 2. Conn E.E. and stumpf, G. Bruenning, R.H. Boi (1987), Outline of Biochemistry by John Wiley & Sons, New York 3. David Rawn, J, (Ed.), (1989), Biochemistry Neil Patterson Publishers 4. Donald and Judith Voet (2005), 2nd edition, J.Niley & Sons, Biochemistry 5. Hall, D.O and K.K.Rao (Eds), (1999),Photosynthesis; 6th Ed., Cambridge University Press. 6. Jocelyn Dow, Lyndsay Gordon, and Jim Morrison, Biochemistry: Molecules, cells and the body 7. Lars Garby and Paul S Larsen (Eds), (1995), Bioenergetics and its foundation; Cambridge University Press. 8. Lehninger et al., (Eds), (1997), Principles of Biochemistry; 2nd ed., Worth Publishers. 9. Mathews, van Holde, and Ahern, (1995), 2nd edition, Biochemistry (Companion Web Site) with 28 chapters. Benjamin/Cummings Publishing Company, Inc., 10. Peter R Bergethon (Ed), (1998), The Physical Basis of Biochemistry; Springer Verlag. 11. Thomas Devlin (Ed),( 2002),Biochemistry with clinical correlations; Wiley-Liss. 12. Tiaz and Zeiger, (2003). Plant Physiology, 3rd edition, Lincoln Taiz and Eduardo Zugier, Parima Publishing Corporation, New Delhi 13. Voet, D and Voet, J.G. (Eds.), (1999), Biochemistry;3rd ed., John Wiley and Sons.

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MLB 302: MOLECULAR BIOLOGY – I 52 hrs Unit I: Concept of Gene: Kinds of genes, gene numbers, functional genes, crypticgenes, pseudo genes, processed genes, overlapping genes.. Gene structure: Structural organization of genes in prokaryotes and eukaryotes-regulatory elements of the genes (proximal or internal, including promoter, operator, activator and enhancers), coding and terminal regions of the gene. 6hrs Unit II: Prokaryotic Gene Expression: Transcriptional Apparatus: RNA polymerase structure, function; sigma factors and their role; and the mechanism of transcription, initiation, elongation and termination. Genetic regulation of sporulation in B.subtilis, role of sigma factors in sporulation. Gene regulation in prokaryotes: Lac operon, Tryptophan operon and Histidine operon -mechanism Genetic regulation in lambda phage: lytic and lysogenic pathway, co-repressor, transcriptional terminators and antiterminators, expression and regulation of early and late genes, site specific recombination. 8hrs Unit III: Eukaryotic Gene Expression: Characteristic features of RNA polymerases - RNA polymerase-I, II and III. RNAP-I promoter: rRNA gene clustering, structural organization. Regulatory region (core sequences and upstream control elements), coding and terminal regions, Role of transcriptional factors (TFs). Mechanism of transcription-initiation, elongation and termination. 7hrs Unit IV: RNAP-II promoter: Structural organization of regulatory, coding and terminal regions of house keeping genes; Genes in response to stimuli-light, chemicals and hormones, and genes regulated in stage and tissue specific manner. Characterization of TATA box, upstream elements, InR elements, enhancer elements, activator elements, response elements, silencer elements/repressor elements; downstream InR elements their position, structure and their function. Transcriptional factors (TFs), activating factors, enhancer proteins and repressors and RE binding factors and their DNA binding sequence elements and composition of factors in different tissue types- general TFs, special TFs, activators and enhancers, repressors/ silencers, mediator complexes response element binding factors.

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Concept of activators, activator domains, co activators and mediators involved in gene expression. 9hrs Unit V: DNA binding proteins- Helix turn Helix, Helix loop helix,, Helix turn beta, Zinc finger, leucine zipper , homeodomains, beta barrels, bZIP and bHLH domains, mode of their binding and regulation of gene expression. 5hrs Unit VI: RNAP-III promoter: Regulatory elements, (internal promoters), coding and terminal regions of 7sLRNA gene, tRNA genes, and 5SrRNA genes; Enzyme, transcriptional factors and assembly of TFS and the mechanism of transcription. 4hrs Unit VII: Regulation of gene expression at transcriptional level : Modes of regulation- Negative and positive type and silencer type of regulation. H2B1Histone gene in sea urchin, globin gene, Gal gene regulation in yeast cells, Interferon regulated gene in animal cells, hormone regulated gene expression in animal cells, light regulated gene expression in plants; silencing of gene activity in Wilmes tumor and yeast mating types . GA induced, auxin induced, ethylene and abscissin induced gene expression. 6hrs Unit VIII: Gene expression and Chromosome remodeling: Structural remodeling during and after transcription; effect of Histones on transcription of class II genes, changes in nuclear positioning, Dnase-1 mapping, Histone acetylation-deacetylation, methylation and demethylation, phosphorylation and dephosphorylation. Role of SW1/SNF and NURFs in remodeling of chromosomes. Organization in the salivary gland chromosomes of Drosophila and lampbrush chromosomes of Xenopus laevis. Role of chromosomal remodeling complexes. 7hrs References : 1. Benjamin Lewin (2004), Gene VIII, Published by Pearson Prints Hall, Pearson Education inc.Upper saddle River, New Jerssey-07458 2. Bruce Alberts, Julian Lewis, Alexander Johnson, J. Lewis, M. Raff (1994), Molecular Biology of the Cell, Garland Publisher Inc., New York 3. Buchnan and Grussem et al, (2000) Biochemistry and Molecular biology of Plants by 5th edition, Oxford University Campus 4. Buchnan B.B., W,Gruissem et al and R.L.Jones (2004) Biochemistry and Molecular biology of Plants by I.K. Internationla Pvt., Ltd., New Delhi 5. Cooper, G M The cell : A molecular approach. 2nd edition, (2000), ASM Press, Washington

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6. Eduardo Diego Patricio De Robertis, EMF De Robertis (1988), Cell and molecular biology, International Ed. Inst. Med. Ltd 7. Gerald Karp (2003), Cell and Molecular Biology, 3rd edition, John Wiley & Sons Publishers.(Concepts and Experiments) 8. Glick B.R. & J.J.Pasternal, (1994), ASM Press, Washington,D.C.Molecular Biotechnology 9. Gupta, PK, (2004) Biotechnology and Genomics, Rastogi Publishers, Meerut 10. Gurbachan S. Miglani(1998), Dictionary of Plant Genetics and Molecular Biology- 348 pages 11. John Marsten Walker, Ralph Rapley (2000), Molecular Biology and Biotechnology

MLB 303 : MICROBIAL BIOTECHNOLOGY 52hrs Unit I: Use of microbes in industry and agriculture: Production of organic compounds by fermentation- Latest biotech methods employed in the production of Ethanol, Acetone and Butanol, Gluconic acid, antibiotics and Enzymes; Methods and protocols for each of the mentioned products: Shake flask method, Bioreactor method, solid state fermentation method, aerobic and anaerobic fermentation method, immobilized cell bioreactor method; Media used, culturing under optimal conditions; Isolation and maintenance of microbial strains, and genetic improvement of strains by genetic engineering and mutation modes: down stream processing-purification of products by adsorption chromatography, affinity chromatography, freeze drying, and in situ recovery methods. 8hrs Unit II: Biotransformation methods: Biotransformation of D-Sorbitol to L-Sorbose, Biotransformation of antibiotics, steroids and sterols. Microbes used for recovery of metals and land reclammation from toxic wastes and chemicals, Bio-insecticides, Bioherbicides, Biofertilizers. Employment of genetically engineered microbes (GEMS): Expression of heterologous genes- completing a partial pathway giving new product. Employing transfer of entire set of genes controlling entire path way of metabolism or creating new products and new reactants, redirecting the metabolic pathway, advantages and disadvantages and limitations. 6hrs Unit III: Commercial scale production by recombinant microbes:

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Production of Restriction enzymes, DNA modifying enzymes- down stream processing methods, Synthesis of Ascorbic acid, Synthesis of Indigo, Synthesis of Amino acids, Synthesis and improvement of production of common and novel Antibiotics, Synthesis of polypeptide antibiotics, Synthesis of Biopolymers-Xanthum gum, Melanin, Animal adhesives; Synthesis of Rubber products. Large scale production of proteins by recombinant microbes: Principles of fermentation cultures, High density cultures. Use of bioreactors for large scale fermentation, harvesting, cell disruption and downstream processing. 7hrs Unit IV: Production of enzymes: Sources and Principles and use of enzyme reactors: Cell culture, large scale production and purification of the products; quality control. Extreme ezymes, Bispecifc enzymes, Ribozymes and Endozymes. Enzyme immobilization and its application: In detergent industry, leather industry, wool industry. Production of glucose from Cellulose; application in food , dairy, beverage and medicinal industry. 6hrs Unit V: Production of single cell proteins: Large scale production and application. Spirulina maxima (Cyanobacteria), Kluyveromyces fragilis (yeast), Candida lipolytica (yeast), Chaetomium cellulolyticum (fungus), Methylophilus methylotrophus (bacterium); Yeast expression systems for production of therapeutical agents: Hepatitis-B surface antigen, Hirudin. Human platelet derived growth factor B, Bovine Lysozyme C2. 6hrs Unit VI: Bioengineering of proteins and its application: Outline of bioengineering of macromolecules a multidisciplinary approach; site directed mutagenesis and computer aided molecular modeling, Steps involved in protein engineering and protein modeling to the desired needs, such as in vitro mutagenesis or synthesis of entire gene, multienzyme with bi or poly functions by gene fusion, chemical modification of existing enzymes; enzymes with characters such as thermo stable, work under non-aqueous solvents, increased Vmax, low Km and high specificity, tolerant to bleach and detergents. 8hrs Unit VII: Development of immunotoxins as magic bullets: Use of Ricin a plant toxin as immunotoxins; drug designing for blocking enzyme activity, blocking receptors from binding to target molecules, for blocking nucleic acid synthesis in cancer or specific cell types. 5hrs

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Unit VIII Environmental Biotechnology: Brief account of pollution control-use of cleaner technology, treatment of industrial effluents, toxic sites and land restoration through alternate aforestation, Mycorhizae,; Bioremediation- cleaning of land and water of oil herbicides, biopesticides by bioengineered microbes, solarization and compatible Biofertilizers, and restoration, conservation of biodiversity-by in situ and ex situ conservation technology, species conservation,, environment and energy- use of alternate fuel sources, use of renewable sources such as biomass, fuel generating microbes. 6hrs

References : 1. 2. 3. 4.

Atlas R.M. (1998) Microbiology : Fundamental and application (IIeds) Mac millan Publishing company Bruijin et al ., (1998). Bacterial genomes, Chapman and Hill Dale J.W. (1994). Molecular genetics and Bacteria. John Wiley and sons Hayes W. (1970). The genetics of Bacteria and their viruses. The English Book society of Blackwell Scientific Publication, Oxford 5. Glick, Molecular Biotechnology , MSM pub, B.R. Glick & J.J.Pasternak, (1994), ASM Press Washington, D.C. 6. Hunderson et al., (1999). Cellular Microbiology Wiley 7. Lewin B (2002). Genes VIII, Oxford 8. Prescot L.M., Hanley, J.P. and Klein, D.A. (1999). Microbiology, WCB Mc Graw Hill , Con MY 9. Roger L.P., John T., Knowler and Daviol P. Leadr. (1992). The Biochemistry of Nucleic acids, 11th edition. Chapmann and Hall 10. Samuel Singer (2001). Experiments in Applied Microbiology, Academic Press New York. 11. Stnely R. Maloy, John E. Cronan, Jr., David Freifelour (1994). Microbial genetics. Jones and Barlett Pub. Bosten. 12. Sullia S.B. and S.Shantharam (1998). General Microbiology , Oxford IBH Publishing Con, New Delhi.

MLB 304: MOLECULAR BIOLOGY OF DEVELOPMENT 52 hrs Unit I:

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Plant System: Biochemical and molecular basis of Growth and differentiation; Concept of growth and differentiation vs. morphogenesis; Site and cell types involved in growth and differentiation. Kinetics of growth, Spatial and material basis of growth and growth trajectory. Polarity fixation: A brief account of polarity development in a fertilized egg cell; and determination of polarity of apices in plant systems - Red algae. 6hrs Unit II Genetic basis: Identity of genes that control development in Arabidopsis; stages of development from embryo, axial pattern, apical basal pattern, radial pattern, tissue identity shoot promeristem, and requirement of gene expression for the development of the above structure in Arabidopsis. The role of Homeobox genes (concept of homeobox genes vs. homeodomain proteins). Interaction of Phytohomones: Interaction of auxins with cytokinins in inducing shoot formation, inhibition of Auxin induced new root formation by cytokinins, Inhibition of GA inducted expression of genes by Abscisic acid, synergistic effects of Auxin and Ethylene, synergistic effects of Auxins and Cytokinins, synergistic effects of Ethylene and Abscisin, overall effects of Phytohormones on each other and their effects on plant development. 8hrs Unit III: Flowering: Characters of shoot meristems that change into floral meristems and development of four types of floral organs, their related genes. Changes in shoot apex, phase changes, Photoperiodism, vernalization effect on growth and flowering. Phytochrome induced responses: Biochemistry of phytochrome, its reponse to light and it effect on plant and floral development. Phytochrom induce responses: Phytochrome induced whole plant responses, biological clock, role of phytochrome in daily circadian rhythm. Phytochrome and gene expression: Different responses of gene expression to Phytochrome A and Phytochrome B, Phytochrome regulation of gene expression- role of cis-acting elements, response elements and trans-acting factors; mechanism of action through multiple signaling pathways; Role of cryptochromes, inhibitor genes of Photo morphogenesis, such as DET and COP genes. Biochemistry of signaling pathway of flowering, identification of hypothetical florigin, Chemical basis of flowering substance, site of synthesis, transport, distribution and site of action and mechanism of action. 10hrs Unit IV: Molecular genetics of flowering: a general account of genes that regulate floral organ development; Arabidopsis as model system , influence of hormones and photoperiod on floral development; Floral organ identity genes, cadastral genes and meristems identity genes, FLORICULA,APETALA (APA 1, APA 2, APETALA3), leafy (LFY), EMF ,DEFICIENS and AGAMOUS genes. Homeotic mutations and their effect on floral organs. Role of MADS box genes in floral organ identity. The role of CRY 1 and CRY 2 genes in

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Photoperiodism Identity of Homeotic genes that control organ identity response factor. ABC model of determining floral identity, effect of hormones and photoperiod on floral development. Regulation of juvenility by TOEPOD genes. 8hrs Unit V: Tissue Engineering – Renewal of adult tissues such as epidermal cells, mammary gland cells, photoreceptor cells in Retina, Gutintestine cells, Alveolar cells in lungs, Liver cells, Blood-endothelial cells, Muscle cells,versatile none cells and fibroblast multipotent cells, stem cells from basal lamina; application and prospects. 5hrs Unit VI: Caenorhabditis elegans: Genes and gene products involved in development. Development from zygote to adult, cell lineage, dauer larval stages. Programmed cell death during development, development of transgenic worms, use of gene knock out experiments to understand the mechanism and its effects. Control of cell lineage –role of Lin genes and regulated PCD in development 5hrs Unit VII: Drosophila- Life cycle, Oogenesis- development of oocyte, role of follicle and nurse cells in the programming of the egg cell. Fertilization and the trigger of a cascade of developmental activation, positioning of specific mRNA, role of maternal genes, Gap genes, pair rule genes, segment polarity genes and Homeobox genes during development., structural organization, developmental genes and their regulation, syncytial blastoderm, cellular blastoderm, gastrulation stages, polarity fixation, segmentation , regulation of gene expression . A general account of Homeobox genes and their role in identifying body organs. 6hrs Unit VIII: A brief account of Development of higher systems- from egg to multicellular level and early development stages and the genes involved in development; Mus-musculus as a laboratory model, Use of transgenic mice and gene knock out in understanding developmental pattern of higher animals. 4hrs References : 1. Bruce Alberts, Julian Lewis, Alexander Johnson, J. Lewis, M. Raff (1994), Molecular Biology of the Cell, Garland Publisher Inc., New York 2. Buchnan and Grussem et al, (2000) Biochemistry and Molecular biology of Plants by 5th edition, Oxford University Campus 3. Goodwin and Mercer- CBS, Plant Biochemistry 4. Gupta PK, (2004) Cell and molecular Biology, Rastogi Publications, Meerut 5. Lewin B (2002). Genes VIII, Oxford

43

6. Lodish,H., Ber, A., Zipuoskry, L.S., Matsudaira, P., Bahimore, D and Damell J. (2001) Molecular Biology W.H Freeman G Co. 7. Taiz, L. and Zeiger, E. (2003). Plant Physiology. 3rd edition. Panima Publishing Corporation, New Delhi/Bangalore 8. Taiz, L. and Zeiger, E. (1998). Plant Physiology. 2nd edition. Sinauer Associates, Inc., Publishers, Massachusetts, USA. 9. Wilkins, M.B. (eds.) (1989). Advanced Plant Physiology. Pitman Publishing Ltd., London.

THIRD SEMESTER PRACTICALS MLB 305: BIOCHEMISTRY-II AND MOLECULAR BIOLOGY - I 104 hrs 1. Extraction and analysis of lipids

44

2. Isolation of mitochondria and chloroplasts : Extraction and analysis of DNA and RNA. 3.Assay of mitochondrial enzymes. 4. Estimation of proteins from mitochondria and chloroplasts 5. Construction of expression vector, PCR protocols 6. Use of enzymes for biodegradation of wastes 7. Purification of enzymes by different protocols 8. Seed germination – by RAPD analysis.

MLB 306 : MICROBIAL BIOTECHNOLOGY AND MOLECULAR BIOLOGY OF DEVELOPMENT 104 hrs 1. 2. 3. 4. 5. 6. 7. 8.

Study of working - Fermentor Estimation of fermentation ability under different condiations ( substrate and temperature) Estimation of percentage of alcohol in yeast fermentation Assay of microbial enzymes ( amylase, protease ) Study of gene expression during embryonic development of Drosophila (LacZ ) Isolation of mRNA from Drosophila ( early embryo ) Analysis of protein profile during flowering. Seed germination – protein profile by iso-electrofocusing.

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NOTE Experiments involving dissection and sacrifice of animals are carried out through computer simulations, models and video demonstration. ---------------------------------------------------------------------------------------------------

45

SYLLABUS FOR III AND IV SEMESTER M.Sc., MOLECULAR BIOLOGY SCHEME OF STUDY AND EXAMINATION SEMESTER III (Hrs / week)

Paper Code

Paper

MLB301

I

Biochemistry – II

MLB302

II

Molecular Biology – I

MLB303 MLB304 MLB305 MLB306

Practical

4 4

--

Title of the Paper

Microbial Biotechnology Molecular Biology of IV Development Pract.I Of paper I & II Pract.II Of paper III & IV III

Theory

Total No. of Hrs/ Semester

Examination

52

Duration (Hrs) 3

Max. Marks 80

Continuous Evaluation 20*

Total Marks 100

--

52

3

80

20*

100

--

52

3

80

20*

100

4

--

52

3

80

20*

100

---

8 8

104 104

4 4

40 40

4

10** 10** Total Marks

50 50 500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical record + 5 marks for visit to Industries/Laboratories

46

SCHEME OF STUDY AND EXAMINATION SEMESTER IV (Hrs / week)

Paper Code

Paper

MLB401 MLB402 MLB403

I II III

MLB404 MLB405 MLB406

Theory

Practical

Total No. of Hrs/ Semester

4 4 4

----

4 ---

Title of the Paper

Immunology Molecular Biology – II Genomics and Proteomics Plant & Animal IV Biotechnology Pract.I Of paper I & II Pract.II Of paper III & IV

Examination

52 52 52

Duration (Hrs) 3 3 3

Max. Marks 80 80 80

Continuous Evaluation 20* 20* 20*

Total Marks 100 100 100

--

52

3

80

20*

100

8 8

104 104

4 4

40 40

10** 10** Total Marks

50 50 500

* 5 marks for Test + 5 marks for Assignment + 5 marks for Seminar +5 marks for Attendance. ** 5 marks for Practical record + 5 marks for visit to Industries/Laboratories

2. TOUR: Tour marks should be considered under Continuous Evaluation. a) There will be a Compulsory Industries / Laboratories visit in all the semesters as per relevance to the subject. b) There will be a compulsory Tour 5 to 6 days to any of the industries outside Bangalore in IV Semester.

47

FOURTH SEMESTER MLB 401: IMMUNOLOGY 52 hrs Unit I: Types of immunity: (a) Nonspecific (Innate) immunity: Anatomic, Physiologic, Phagocytic barriers (b) Specific (Adaptive) immunity: Active immunity Naturally and artificially acquired; Passive immunity - Naturally and artificially acquired and Adoptive immunity (c) Brief description of cell mediated immunity and humoral immunity 5hrs Unit II: Organs of the immune system: (a) Primary lymphoid organs: Bone marrow, Thymus (b) Secondary lymphoid organs: Spleen, Lymph nodes, Mucosalassociated lymphoid tissue. 5hrs Unit III: Cells of the immune system: (a) Hematopoesis (b) Lymphoid cells – B Lymphocytes, T Lymphocytes (TH, TC and TReg cells), NK Cells (c) Macrophages (d) Granulocytes (e) Mast cells (f) Dendritic cells (g) Clinical uses of hematopoietic stem cells Maturation, Activation and Differentiation of T and B Lymphocytes, Antigen processing and Presentation: Processing and presentation of endogenous and exogenous antigens, presentation of non-peptide antigens 8hrs Unit IV: Antigens: Immunogenicity and Antigenicity, Factors influencing immunogenicity, Epitopes, Haptens 4hrs Unit V: Antibodies: (a) Classes of antibodies – IgA, IgD, IgE, IgG and IgM (b) Structure of IgG (c) Cellular kinetics of antibody synthesis (d) Organization and expression of Ig genes - l - Chain multigene family, k - multigene family, Heavy chain multigene family, V-J rearrangements of light chain, V-D-J rearrangements of heavy chain, organization of Recombination Signal Sequences (RSSs). (e) Other mechanisms causing antibody diversity – Junctional flexibility, P-addition, Naddition, Somatic hypermutation, Combinatorial association of light and heavy chains (f) Class switching (g) Monoclonal antibodies: Formation and selection of hybridomas, Production and Clinical importance of monoclonal antibodies 10hrs 48

Unit VI: Major Histocompatibility complex: Structure of Class I and Class II MHC molecules, Polymorphism of peptide binding region, Class I MHC – peptide interaction, Class II MHC – peptide interaction 4hrs Unit VII: Vaccines: Primary and Secondary responses, active and passive immunization, Whole organism vaccines, Purified macromolecules as vaccines, RecombinantVector vaccines, DNA vaccines 3hrs

Unit VIII: Hypersensitivity: IgE mediated hypersensitivity (Type I), Antibody mediated cytotoxic hypersensitivity, Immune complex mediated hypersensitivity (Type III), Delayed type hypersensitivity (Type IV) 5 hrs Immunodeficiencies: Primary immunodeficiencies – Lymphoid immunodeficiencies, Myeloid immunodeficiencies, Secondary immunodeficiencies (AIDS) 4hrs Autoimmune diseases: Autoimmune haemolytic anaemia, Hashimoto’s disease, Grave’s disease, Myasthenia gravis, Pernicious anaemia, Systemic Lupus Erythematosus 4 hrs References: 1. Abul Abbas, Andrew Lichtman, and Jordan Pober, (2005), Cellular and molecular immunology, Saunders Publishers, 5th edition, 576 pages plus CD 2. Abul Abbas, Saunders,(2006), Basic Immunology, Updated Edition 2006-2007 (Paperback) by Publishers; 2nd edition 336 pages 3. Ashi K Chakravarty, (2006), Immunology and Immunotechnology , Ist edition, Oxford Press. 4. Charles Janeway, Jr. and Paul Travers, (2004), Immunobiology the immune system in health and disease, by. Garland Science; 6 edition, 800 pages 49

5. Gupta P K, (2004) Cell and Molecular Biology, Rastogi Publications, Meerut 6. Ivan Roitt, Jonathan Brostoff, and David Male. Mosby, (2006), Immunology, London. 7th edition, 544 pages 7. Lodish et al., (2001) Molecular Biology, W.H.Freeman G Co. 47 8. Thomas Kindt, Barbara Osborne and Richard Goldsby, (2006), Kuby Immunology. W. H. Freeman & Co., Sixth edition, 2006 9. William E Paul, Lippincott Williams & Wilkins;(2003) Fundamental Immunology (Hardcover) by 5th Bk&Cdr edition ,1502 pages

MLB: 402 MOLECULAR BIOLOGY - II Unit I: 52 hrs Control of gene expression at Post transcriptional level: 1. Processing of rRNA: precursor rRNAs, types and structural and functional features of sno-RNAs and sno-RNPs and their role in splicing rRNAs. Processing of pre-tRNAs: size of pre-tRNAs, the number, size and position of tRNA introns; two types of splicing and the mechanism of splicing. Pre-mRNAs processing: Characteristic features of pre-mRNAs or Hn RNAs, structure and sizes of hn RNAs; hnRNP proteins, mRNP proteins; introns and exons and their characters; Processing of pre mRNAs; Capping and polyadenylation: Enzymes and the time of capping, mechanism of capping, its importance. Factors, site, enzymes and the mechanism involved in Poly(A) addition, importance of poly(A) tail; poly(A) binding proteins and their role. Splicing: Characteristic features of introns splice junction site and intron’s internal sites, splicing signals and signal sites; SnRNAs, their structural and functional features and their associated proteins; Mechanism of splicing event, the role of specific snRNA and SnRNPs; spliceosomal assembly and the mechanism of splicing. Processing of Histone mRNA and the role of sn-U7 RNA. 8 hrs Unit II: Alternate splicing: Concept of alternate splicing and its implications. Alternate splicing of Fibronectins, collagens, Example from Dscam splicing in Drosophila, alternate splicing of vertebrate inner ear cochlea for registering sound waves, A 50

brief description of alternate splicing in sex determination of Drosophila. Alternate splicing in mouse/human immunoglobulin u-heavy chain mRNA. Trans splicing: trans-splicing in C.elegans and Trypanosome pre-mRNAs, and some Adenoviral mRNAs, where the leader sequence is same in 25% or more of the mRNAs; splicing components and SL-RNA and other snRNA-RNPs involved in the mechanism of transplicing. Pre-mRNA Editing: Editing Apo-lipoprotein mRNA and Glutamine receptor mRNA, features and mechanism. Special features of few mitochondrial faulty pre-mRNAs (called pre-edited mRNAs) in Trypanosomes and Leishmania; editosomes, and characters and their composition, genes for Guide RNA and the mechanism of editing. Self-splicing introns: Group-I introns, Group-II introns, Group III introns, Twinintrons: their characters and functions; Informosomes: Stored mRNAs in mature egg cells , normal cells and seeds, role of mRNPs, importance of poly(A) size, polyadenylation signal elements, reactivation of mRNAs by reactivation of Poly(A) addition and its regulation. 10 hrs Unit III: mRNA transport: Structures and proteins involved in the transport of rRNA, tRNA and mRNAs; mechanism of transport. mRNA stability and turn over: sequence elements found in the 5’ leader sequences and 3’ non-coding regions and their structural features, relationship between such sequences and sequence derived structures and stability; mechanism of protection and the mechanism of degradation and causes; ex. Casein mRNA, Transferrin mRNA, Ferritin mRNA.. 8 hrs Unit IV: Control of gene expression : Prokaryotic translation Translation apparatus; ribosomal subunits, initiator-tRNAs aminoacyl-tRNAs, initiating factors, elongation factors, termination factors, and their characteristic features, mechanism of chain initiation, elongation and termination; production of specific proteins on translation of a polycistronic mRNA. Mechanism of translation, Post translational processing of polycistronic polypeptides, and targeting the protein to periplasmic space or to the membrane. Regulation of protein synthesis, autogenous regulation, stringent response type regulation. 6 hrs Unit V: 51

Eukaryotic translation: Translational apparatus- ribosomes, initiator-tRNAs, aatRNAs, initiation factors, elongation factors and termination factors, their composition and functions; mechanism of translation; Regulation of protein synthesis: Regulation of translation at mRNA level, regulation at chain initiation factor level, ex. Heme regulated translation, regulation of Ferritin synthesis, and Transferrin receptor synthesis and interferon mediated regulation. Membrane free site-actin filament associated synthesis, localized synthesis, synthesis on ER. 6 hrs Unit VI: Post translational processing: Co translational processing- transferring the translating system onto ER and transferring the protein into the lumen of ER, the role of SRP particles, docking proteins, and signal sequences in targeting the protein and also in orienting the N and C- terminal ends of the proteins. Mechanism of transfer of proteins into ER lumen. 5 hrs Unit VII: Post translational modification and targeting: Processing of proteins in the lumen of ER, transfer of the same into cis surface of the Golgi membrane, further processing, and transferring to mid and trans-golgi membranes, modification such as glycosylation, sorting and packing and delivery of the same to specific destination, factors and the proteins involved and the mechanism. Transfer of membrane free synthesized proteins into organelles such as nucleus, chloroplasts, mitochondria and glyoxysomes; the signal sequences and the enzymes and factors involved in the mechanism of transfer and targeting. Processing of Pre-pro-proteins: Regulated cleavage of polyproteins and pre-pro proteins in stage specific and tissue specific manner. Splicing of proteins, Removal of introns and joining of exons- its mechanism. 6 hrs Unit VIII: Protein stability and turnover: Sequence based structural form, unstable proteins, protein degradation, and ubiquitination of condemned proteins and degradation by proteosome; structure and features of Proteosomes and the mechanism of degradation. 3hrs

References: 52

1. Buchnan, B.B. and Wilhelm Grussem et al., (2000) Biochemistry and Molecular biology of Plants , American Society of Plant Physiologists, Rock Ville, USA, Maryland 2. Eduardo Diego Patricio De Robertis, EMF De Robertis (1980) Cell and molecular biology 3. Gerald Karp, (1996) Cell and Molecular Biology – Concepts and Experiments. John Wiley and Sons, Inc., New York 4. Gurbachan S. Miglani (1998), Molecular Biology:Dictionary of Plant Genetics and Molecular Biology, - 348 pages 5. James Darnell, Harvey Lodish, Paul Matsudaira, Arnold Berk, S. Lawrence Zipursky, (1998) Molecular Biology of the cell 6. John Marsten Walker, Ralph Rapley (2000), Molecular Biology and Biotechnology 7. Russell L. Jones, Virginia (EDT) Walbot (1995), Annual Review of Plant Physiology and Plant Molecular Biology - Page 22 8. Sambrook,J; Russel,D.W., (2001)Molecular Cloning: A Laboratory manual, cold spring Harbor Laboratory press, cold spring Harbour, New York 9. William H Elliott, Daphne C Elliott (1997), Biochemistry and molecular biology

MLB403: GENOMICS AND PROTEOMICS 52 hrs Unit I Sequencing: Use of large scale sequencing and application in identifying species, varieties and relate phylogeny. Physical map: Use bottom-up approach or Top down approach; Sorting out chromosomes by Fluorescence activated chromosome sorting method (FACS), Use of PFGE-Pulse field gel electrophoresis or CHEF-Counter clamped homogenous electrical field electrophoresis, Use of molecular markers, use of chromosomal aberrations such as deletions and fragmentation, preparation of YAC libraries or Bacmid libraries, RE mapping, chromosomal walking and jumping, contigs, use of in situ hybridization (ISH) and FISH, use of EST tags (expressed sequence tags) and STS tags (sequence tagged sites) use of positional cloning of cloned sequence of a chromosome, and candidate gene approach, developing high resolution mapping, 53

identification of genomic sizes by Resolution-Gap mode. Development of integrated genomic map using all methods. 8hrs Unit II: Functional Genomics: Transcriptomes-overall transcripts of a given tissue and tissues; use of Northern blot, dot blot and reverse northern blot, use of subtractive library, additive library, RNase protection and RT-PCR techniques; techniques for analysis of steady state gene expressions by EST tags and CDNA libraries, characterization of each of them and relating each of them to each kind in terms of nucleotide sequence to their amino acid sequence; detection of ORFs, gene knock outs, construction of two expression plasmid systems in one cell to find out gene interaction. 8hrs Unit III: DNA Micro array techniques-spotted arrays, printing techniques and spotted array techniques, difference between spotted arrays and Oligochips; Transcriptomics and data processing. Construction of DNA-chips to find out regulation of gene expression and to detect mutations. Gene expression measurement through DNA micro arrays and SAGE , Protein levels through twodimensional gel electrophoresis and mass spectrometry, including phosphoproteomics and other methods to detect chemically modified proteins. 6hrs Unit IV: Comparative Genomics: Concept of orthologs and paralogs in gene evolution, protein evolution through exon shuffling, comparative genomics of bacteria and large microbial genomes, comparative genomics of closely related bacteria in particular and microbes in general, comparative genomics and physiological phenomena,; comparative genomics of organelles, comparative genomics of eukaryotes to identify genes and regulatory elements, evolution of key proteins, and evolution of species; comparative genomics and molecular mechanism to generate new gene structures. 8hrs Unit V: Proteomics: Concepts, Protein array from a given tissue or tissues, detection and screening techniques. Use of 2-D PAGE, sensitivity and resolution and representation of 2-D gels, multiplexed analysis to show expression profiles,; use of multidimensional liquid chromatography, Mass spectrometry and high throughput protein annotation, Matrix assisted laser desorption (MALDI), Electrospary ionization (ESI),TIME flight (TOF), collision induced dissociation (CID), special strategies for qualitative and quantitative analysis. 54

Protein array for expressional analysis, profiling and functional analysis; antibody array for the capture of specific protein, antigen array to measure antibodies in a solution, use of BIOchips. Structural Proteomics-use of X-ray diffraction crystallography and NMR, circular dichroism (CD), use of algorithms for comparisons, functional assignment. 8hrs Unit VI: Protein-protein interactions: genetic approach, use of comparative genomics, use of library based screening and analyzing interactions, invitro expression libraries, yeast two hybrid system,, matrix approach,, analysis of protein complexes by affinity purification and MS, protein localization in a cell. 5hrs Unit VII: Metabolomics and metabolic engineering: Concepts, methods used and application. Cloning and expression of heterologous genes to change metabolic pathway to the desired product or to improve the desired metabolic product; altering feed back inhibition, altering the regulation of a desired metabolic pathways.. Metabolomics-for small-molecule interactomics for interactome 4hrs Unit VIII:

metabolites,

glycomics

for

sugars;

Systems Biology; Markup Language; List of omics topics in biology; Gene regulatory network; Metabolic network modeling ; Model Computer simulation; Important publications in systems biology Systems theory; Systems ecology ; Regulome; Biomedical cybernetics; Different methods for analyzing metabolites. Choosing analytical methods to the need of an organism. Basic mass spectrometry-use of Gas (GC/MS and liquid chromatograph (LC/MS). Use of Tandem Mass spectrometry, sample selection and sample handling, use of data base for preparing global biochemical networking. 5hrs References: 1. Brown, T.A., Genomes (1999). John Wiley &Sons 2. Daniel, C. Leibler, (2002). Introduction to Proteomics: tools for new biology, Human Press, Totowa, NJ 55

3. Dennis, C and Gallaghar, R (2001). The human genome. Nature publishing group, U.K. 4. Kohane IS, AJ Butte, A Kho (2002), Microarrays for an Integrative Genomics - group of 2 », - MIT Press Cambridge, MA 5. Liebler DC (2002), Introduction to Proteomics: Tools for the New Biology - group of 12 » 6. Liu, BH (1998), Statistical Genomics: Linkage, Mapping, and QTL Analysis - books. 7. Maleolm and Goodship (2001). Genotype to Phenotype. 2nd ed. Bios Scientific publishers. 8. Palzkill, Timothy : (2002) Proteomics, Kluer Academic Publishers 9. Pechkova E, Nicolini, C (2003), Proteomics and Nanocrystallography 10. Pennington, S; M.J. Dunn (eds) (2001) Proteomics: from protein sequences to function. Springer Publications. 11. Puhler,A(1993), Genetic engineering of microorganisms, WCH Germany 12. Saito K, RA Dixon, L Willmitzer (2006), Plant Metabolomics - group of 2 » 13. Sehena,M (1999), DNA microarrays: A Practical approach, Oxford University Press, Oxford 14. Simpson RJ, JL Hotchkiss (2003), Proteins and Proteomics: A Laboratory Manual 15. Strachen,T Read, AP. (1999), Human Molecular Genetics, 2nd edition, John Wiley & Sons. 16. Tomita M, T Nishioka , lavoisier (2005), Metabolomics: The Frontier of Systems Biology - group of 3 » 17. Tomita, Masaru (Editor ), Takaai, Nishioka (Editor ), Metabolomics 18. Wilkins MR, RD Appel, DF Hochstrasser, KI Williams (1997), Proteome Research: New Frontiers in Functional Genomics., Springer –verlag, New York 19. Zhou, T, Thomas, D.K.,Y.XU and Tiedge, J.M. and Wiley Liss (2004), Microbial functional genomics

MLB 404: PLANT AND ANIMAL BIOTECHNOLOGY PART A : PLANT BIOTECHNOLOGY 52 hrs 56

Unit I Principles of Tissue culture: Preparation of media, its components, autoclaving principles, preparation of explants and culturing. Use of Phytohormones: Understanding of Phytohormones and their effects on plant morphogenesis, interaction of the same in morphogenesis. Strategies to induce callus and plantlets from different types of explants. Culture Types: Methods of embryo culture and embryo rescue, Meristem culture, virus free plants, ovule culture, endosperm culture. Large scale Micropropogation, hardening and its application. Selection for somaclonal variants and creation of somoclonal variants. 6hrs Unit II: Cryopreservation: Germplasm sources and techniques of Cryopreservation Haploid plants: Use and mode of ovule and anther culture for haploid plants, application of Haploids in agriculture. Protoplast culture: Preparation of protoplasts and culturing technique, modes of protoplast fusion to produce cell hybrids and cybrids, chromosomal stability; its application. Liquid culture: Production of unicellular cell population by liquid culture and application liquid culture in creating embryoids, mutants. Transgenic plants and Large scale tissue culture: Kinds of culture: agar cultures, suspension cultures, Batch cultures, continuous and Multistage Bioreactors, immobilized bioreactors; Improving and enhancing yield, development of high yielding clones, and use of elicitors. Large scale culture: Single cell, Solid callus and Liquid culture, maintenance of the liquid culture, extraction methods and scale up of the process-for secondary metabolites; production of single cell proteins(SCPs). 8hrs Unit III: Transgenic Plants: Plant cell culture for biotransformation. Biochemical production of Agrochemicals, Medicinal, Cosmetic and food additives, Enzymes, Plantibodies and vaccines, Biopolymers and vitamins.Vectors for plant transformation- Binary vectors and Integration vectorsits characteristic features in detail. Construction of a gene in expression mode Use of selection marker gene and how to get rid of the harmful antibiotic or toxin genes Use of terminator technology. 57

Methods of gene transfer: Electroporation, Gene gun and Agrobacterium mediated. Ti and T-DNA, mechanism of DNA transfer and integration, design of vectors on the basis of Ti-plasmids.Tissue regeneration and screening- methods . Organelle Engineering :Construction of vectors with proper regulatory elements. Transfer of genetically engineered DNA clones into chloroplasts of Chlamydomonas, and higher plants.. Targeting cytoplasmically made proteins into chloroplast. Targeting cytoplsmically made proteins into Mitochondria 8hrs Unit IV: Disease Resistance transgenic plants : Disease Resistant to viruses -capsid gene, antisense to capsid gene, defective transport protein and RNA and RNAi and Ribozyme mediated resistance. Disease Resistant to fungi by engineering Chitinase (b-1, 3-glucanase gene. Disease resistant to bacteria by Lysozyme gene. Resistant to pests- Bt-toxin gene, protease inhibitor genes. Use of biological insecticide (Baculoviruses) and herbicide resistant strains. . Transgenic food crops: Increase in essential amino acids in cereal seed proteins (phaseolin promoter and 2sAlbumin gene; E.coli dihydropicolinate synthase (DHPS gene) with signal sequence for chloroplast, for increasing lysine content. Increase and change in the quality of oils in Brassica species (increase in medium chain fatty acids and converting unsaturated fatty acid to saturated fatty acids). Prevention of discoloration of Food, Increase in sweetness and flavor in fruits. Increase in Amylopectin (by antisense to starch synthase). Improvement in amylase synthesis by ADPG pyrophosphorylase. Plants as Bioreactors: Production of plantibodies, Production of vaccines. Production of biopolymers. Production of water soluble oleosins. 8hrs PART B: ANIMAL BIOTECHNOLOGY Unit V: Methods of animal cell preparation: Protocols used for cell cultures from the tissues and culture methods and maintenance, Large scale animal cell culture for commercial production of specific proteins, Igs, Interferons, vaccines, Monoclonal Antibodies, Hybridoma cells and other down stream processes. Animal tissue culture: skin cultures, Neuronal cell cultures, muscle cell cultures, Cartilage culture, blastocysts cell culture, whole embryo culture, Stem cell 58

research: use of stem culture methods for transformation and tissue transplantation. Kinds of stem cells and characters and potentials, methods to obtain such cells, culture and maintenance of cells; methods to induce stem cells to differentiate into specific tissues; Application of stem cells in medicine, research and therapy; ethical problems. 6hrs Unit VI: Animal cell Transformation and immortalization: Methods employed for animal cell transformation, viral mode, oncogene mode or genetic manipulation mode, Properties of transformed cells, causes and mechanism of transformation, genes involved in transformation, application of cancer cells in biotechnology. IVF in humans and animals: In vitro fertilization, oocyte culture, in vitro fertilization and embryo transfer (IVF-ET). Design of vectors for genes expression in animal cells; Integrating vectors, episomal high copy number vectors with inducible promoters. Production of recombinant viruses for targeting specific tissues. Methods of transfer: Electroporation, PEG method, and transfection method. 6hrs Unit VII: Transgenic animals: Methods employed in introducing cloned genes; protocols for developing transgenic animals; use of fertilized egg cells, use of blastocyst cells; success and failures, problems. Transgenic sheep, goat, fishes, cattle, mice, pigs and birds. Applications and ethical issues. Animal cloning: Techniques used in animal cloning- transfer of whole 2n nuclei to enucleated cells (ex. Xenopus and other frogs), cultured cell fusion, use of embryonic cells, applications and ethics. 5hrs Unit VIII: Human gene therapy: Ex vivo and in vivo gene therapy, viral gene delivery system, non viral delivery system, pro-drug activation therapy, oligonucleotide correction methods. Cell mediated therapy - blocking cell receptors in Hodgkin’s melanoma, treating cancer cells with Tissue specific Tumor infiltering lymphocytes (TILs) and interleukins. Treating transformed hepatocytes for defects in clotting factors and other liver specific defects. Development of vaccines against specific cancer types. Gene therapy for SCID (severe combined immunodeficiency), Antisense RNA as therapy. 59

Cancer therapy: use of specific vaccines, use of antisense or RNA interference. 5hrs References: 1. Altman, A ( 1997), Agricultural Biotechnology 2. Bhojwani SS, MK Razdan (1983), Advanced immunology - group of 2 » Plant Tissue Culture: Theory and Practice, - elsevier.com 3. Freshney RI, JRW Masters, J Masters (2000), Animal Cell Culture: A Practical Approach, 5th edition, Wiley-Liss Publication 4. Goldsby RA, TJ Kindt, BA Osborne (2000), Kuby immunology, academicbooktrade.co.uk 5. Houdebine, LM (2003), Animal Transgenesis and Cloning - group of 3 » 6. Krimsky DS, RP Wrubel (1996), Agricultural Biotechnology and the Environment: Science, Policy, and Social Issues - group of 3 »

FOURTH SEMESTER PRACTICALS MLB 405: IMMUNOLOGY AND MOLECULAR BIOLOGY - II 104 hrs 1. Raising antibodies against BSA in rabbit 2. Ouchterlony double diffusion 3. Protein electrophoresis by using serum from control and antigen immunized rabbits 4. Identification of pathogens by ELISA. (Kit method) 5. Preparation of genomic DNA. 6. PCR amplification of an identified gene. 7. Subcloning of a gene for expression in a prokaryotic expression vector 8. Protein expression analysis MLB 406: GENOMICS AND PROTEOMICS & PLANT AND ANIMAL BIOTECHNOLOGY 104 hrs 1. Study of proteins by native gel electrophoresis 2. Study of proteins by SDS-PAGE 3. Study of proteins by 2 D gel electrophoresis 4. Western blotting. 5. Computational analysis of the proteome of a given organism 6. Transient expression of a cloned gene in animal cells by electroporation and analysis 7. Tissue culture, micropropagation, anther culture 8. Protoplast culture and fusion 60

9. Plant transformation and molecular analysis by PCR ----------------------------------------------------------------------------------------------------

NOTE Experiments involving dissection and sacrifice of animals are carried out through computer simulations, models and video demonstration. ------------------------------------------------------------------------------------------------------

61