Week 8 Full

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f A:ZaraTabi([email protected]) AIM:ztabi06OH:ByemailAppt.

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BICD 120:Dr. Crawford Week 8

TI

Mechanism: -The

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/ \ - OiiOative burst) creation ofRNSandROS) Production F?ru.*a offreeradicals Production of toxins to kiIIEVERYTHING in the area uhetucr p/anr m -*M cz// . Y,;/'rr%i{ ttn* ,,1.2) &4 '94rn ' - Hyrolases fu/hwn T4/, -PhYtoalexins S ,^ '(! ' -- vllu "induces u u u i plant O*idatire cl.lvtr u bunst u lJt -)7 lllu cell l death P l cl l l l ' utrl , prllsVarwoffvts _'' " * "V /

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V . Plant (basal) detects markers/elicitors frompathogen ) lnduces innate immunity Tt4aint' 5| "' - Release ofROS, hydrolases, calose t \ - Whataresome examples ofelicitors? E\ .- FlaglileilNi proteins pruterns(shared Fragments ofueil ul cellwalls, w4il5,uuiluervsu conserved majority byaa rilajonly ofoac., or bac.) tsilareq oy t\ A\ . Pathogens produce proteins may virulence to counteract innate immunity e\ 5,-\ fn nf rria canralinn crrclam e.tu;t^, +^ nnr)4ilA/)-f7btA -- llolirrorarl plant Delivered inside viahrna ---"> f,lnli/a4 totha theincida offha thenlanl typellllll secretion system---> n t\ SlWf#* to\ proteins, virulence theplantproduces R-proteins* zfatlsalue/ rhln 'p:orrru ' Tocounteract t/ proteins tobindtovirulence ) induces hypersensitive - Made response ( \ . Release \ phytdabxins, ROS, RNS, Progammed celldeaih, Increased lignin . ..d, -

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response -Hypersensitive onry initiared .tU, whena plantis -^rl#,iini"KAffi#i"6k-W#"*fn'itti*4/u,/ns - Whathappens notable tofendoffaninfectiontn* , 0( SP"af'tttuJ i'spreadsthroughout Systemic infection vt4ftU?r theplant, t u in $ o e c' *lfr'!Y.,,, AGROBACTERIUM! A Slar'rr SJyprr

Pathoqenic?

A.radiobacter ) noInfection A,rizogenes ) Hairyroottumor A.Tumefaciens ) Undifferentiated tumor/gall

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TheBasic -lnfection: Characteristics! 1)Plants should bewounded toinfect iscrucial too) optimum at12-18 - Timeofinfection hrsafterwounding 2)Temperature ) willnotinfect sensitive athightemperatures planttumor 3)Infected cellscangrowinmedia without hormones ) transformed needpresence togrowuntransformed cells, - REMEMBER: ofhormones inmedia 4)Tumor cellsproduce opines Acidbonded released andAmino together, intothesoil - Sugar asnutrients - OnlyAgrocanusethecompound go/ion'/'/""' ) TiPlasm - Whatisneeded forplantinfection? id -*fitlf d,s////,fut/vt {?

4 Parts of a Ti Plasmid: - Framework ofReplication 1)ORI= Origin plasmid whenbacteria divides toreplicate - Needed genes 2)Opine CATABOLISIM source Opine touseasa nutrition toMetabolize - Needed 3)VirGenes T-DNA andtransfer needed tomobilize Virproteins - Produce . Makess COpy OfT-DNA-,

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a CloserLook 4)TheT-DNA:

) Produces Opines Genes BIOSYNTHETIC - Opine growth undifferentiated for causing ) responsible - Oncogenes = cyctokinin 'tmrl Produce Produce auxin = ' tms1,2= , ,v,4 . Different genes :l,i/^,.v'-' -' aremutated! willoccurifthese results tumor **? ! Fillinthechart: Agrogenes

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Auxin

Growth NoGrowth!

TmrTmsTmr-Tms* + ntot *uue{'L D (Tmr*Tms.9't ot)irlgtl^A {.|(w.t.) Tmr*Tms* dt uvt J tzu,ttt h aVA .t'1.) [^^JL + to media inghormones the cellsbyadding withUN-TRANSFORMED results - Canreproduce (w.saltsandsucrose) . Keytoregenerating/propagatirE plantcellsintissue cultures a- /Ph'y'/lar ,olnr/fto' plant propagation? you of basic steps the - Can describe first? orAuxin incytokinin thecallus - Doyouplace | .+ Ailxrn Znd, 4+ok-rVtrv'r \s* I PlantTransformation plants oftheirchoosing! withgenes totransform canbeusedasa vector realize A.Tumefacians Scientists T-DNA region from oncogenes toremove - Need ryineandESPECIALLY . ^- ,. !.,

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- T-DNAmodifications: . Multiple forgeneofchoice sitesthatallowinsertion Restriction Site(MCS)Cloning . Selectable plantcells thetransformed Todistinguish marker(Kant) genesforKanamycin resistance - Usually // -/,n ,"r4r4u/.,///r t.u/ 4t'tt ;/"5 n7vA7fuatPt/ /'nf marl/f 4hn,ts "' 'n'br*rrO/r; , i

carbon source, with 2)Apply cellstoagarplate[w/salts, plantandapplyAgroculture transformed 1)Wound (aux Kanl. Allow todevelop hormones cyt), and Callus gene & TiPlasmid w/ ofinterest

toagarplate[w/salts,carbon source, 4)Cutshootandtransfer carbon toagarplate[w/salts, callus 3)Transfer (aux Allow root hormones ONLY), andKan]. source, (cytONLY), Allowshoot andKanl. hormones development development

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Process Plantentirely composed ofcells Completed! geneofinterest! containing

ofplantkansformation system -Binary . Useofa deleted plasmid (missing T-DNA) anda T-DNA Tiplasmid

. Because plasmid easier toworkwith! smaller, issignificantly T-DNA -rd// 7lttuudf al t/cg /rrufalt/ '//ae- {fu {

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