Plasmids

Plasmids Schematic drawing of a bacterium with its plasmids. (1) Chromosomal DNA. (2) Plasmids

RB Shahi

 Capable of autonomous replication  i.e. self- replicating  A plasmid is a DNA molecule separate from the chromosomal DNA  i.e.. Extra chromosomal(replicons)  That are commonly present in bacteria, archaea, and yeasts  Typically circular and double-stranded but Linear plasmids have been found in a variety of bacteria,e.g. Streptomyces sp. and Borrelia burgdorferi  Often carry genetic information that is useful only under atypical conditions, and are sometimes lost from the cell (in non-selective environment)

 1952: J. Lederberg reviews the literature on cell

heredity and suggests the term "Plasmid" for all extrachromosomal hereditary determinants.

 1954: Pierre Fredéricq and colleagues show that

colicine (plasmids) (large toxin proteins (50-70kD) ) behave as genetic factors independent of the chromosome.

 1958: François Jacob and Elie Wollman propose the

term "Episome" to describe genetic elements such as F factor, colicine, and phage lambda, which can exist both in association with the chromosome and independent of it.

 By the end of the 1960s, both the genetic and physical nature of plasmids and cytoplasmic heredity had been known in detail and the "Modern Period" of Plasmid Research starts - recombinant DNA technology.

 1970s-80s: the Cytoplasmic mitochondrial and chloroplast DNAs in green algae and plants were continuously being studied and their circular forms of dsDNAs.

Plasmids frequently carry genes associated  Resistance to antibiotics, metal ions and ultraviolet light;  Virulence;  Bacteriocin-production;  Degradation of toxic organic compounds; and  The ability to transfer from one bacterial cell to another.

Conformations of Plasmid DNAs (Plasmid DNA may appear in the following five conformations)

1) "Supercoiled" (or "Covalently Closed-Circular") DNA is fully intact with both strands uncut. 2) "Relaxed Circular" DNA is fully intact, but "relaxed" (supercoils removed). 3) "Supercoiled Denatured" DNA. small quantities occur following excessive alkaline lysis; both strands are uncut but are not correctly paired, resulting in a compacted plasmid form. 4) "Nicked Open-Circular" DNA has one strand cut. 5) "Linearized" DNA has both strands cut at only one site.

Nicked DNAs

Linear DNA

Super Coiled SC Relaxed region

Conformation Vs Electrophoretic mobility The relative electrophoretic mobility (speed) of these DNA conformations in a gel is as follows:

Nicked Open Circular (slowest) Linear Relaxed Circular Supercoiled Denatured Supercoiled (fastest)

Classification of Plasmids • Based upon the number of copies per cell, plasmids are classified into two types. – Stringent plasmids – Relaxed plasmids

• Based upon plasmid incompatibility: inability of two different plasmids to co exist in the same cell in the absence of selection pressure – Compatible – Incompatible

• Based upon the ability to take part in conjugation, plasmids are of two types. – Conjugative – non conjugative

• Host Range – Broad spectrum – Narrow spectrum

• Phenotypic effects – – – –

Fertility Bacteriocinogenic plasmid Resistance plasmid (R factors) Etc.

Stringent plasmids • These plasmids exist in small numbers, i.e. <100 copies/cell (limited number of copies per cell) • Under the control of bacterial genome for replication and segregation • Generally, conjunctive plasmids are mostly stringent plasmids.

Relaxed plasmids • These plasmids exist in large numbers, i.e., > 100 copies/cell.(multiple copies per Cell) • not under the control of bacterial genome for replication and segregation • Generally they are of low molecular weight • most of them are of the non conjugative type.

Compatible • Have different replication control

Incompatible • Have same replication control

Conjugative • those which take part in conjugation • They have tra genes which help in conjugation, • e.g. F. Plasmids

non conjugative • those which does nittake part in conjugation • They lack tra genes which help in conjugation, • e.g. pBR 322

Broad spectrum • Replicate in many types of bacteria – RSF 1010(wide range of Gram-negative and Gram-positive bacteria-non conjugative) – RP4(most Gram-negative bacteria-conjugation) – etc

Narrow spectrum • Replicate in only one type of bacteria – pBR322 in E .coli

Maintenance of Plasmids •

Two factors are important –

One is their faithful replication at least once in each cell cycle: Regulated 1. by antisense RNA and 2. by binding of essential proteins to iterons

–

their accurate partition so that each daughter cell receives at least one copy

Rop=Repressor of Primer

Function of Plasmids 1) Antibiotic resistance 2) Antibiotic production 3) Degradation of aromatic compounds 4) Haemolysin production 5) Sugar fermentation 6) Enterotoxin production 7) Heavy-metal resistance 8) Bacteriocin production 9) Induction of plant tumours 10)Hydrogen sulphide production 11)Host-controlled restriction and modification What are cryptic plasmids?

Based on their function, there are five main classes  Fertility-(F)plasmids: they are capable of conjugation or mating.  Resistance-(R) plasmids: containing antibiotic or drug resistant gene(s). Also known as R-factors, before the nature of plasmids was understood  Col-plasmids: contain genes that code for colicines, proteins that can kill other bacteria.  Degrative plasmids: enable digestion of unusual substances, e.g., toluene or salicylic acid.  Virulence plasmids: turn the bacterium into a pathogen. Hly Plasmid :α-haemolysins of RBCs Plasmids can belong to more than one of these functional groups.