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f A:ZaraTabi(ztabi@ucsd.edu) BICD
120:Dr.
Crawford
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AIM:ztabi06OH:ByemailAppt. ,id .^,t Week
8 . u
Microbiarperense*rfiftrn g{
/ ..,p TI SI
t7 ^*
-2Basic
things
occur: i ,rdt'' ,,il!' 6f N
1)Walling
offof infected
area) How? .Z', ,, '
.L,r,t
ttltt 7 -Ligninproductionincreases
-Ligninproductionincreases
/ / \
\-\
F?ru.*a \ - OiiOative burst) creation ofRNSandROS) Production offreeradicals \
Production of toxins to kiIIEVERYTHING in thearea uhetucr p/anr m -*M N
&4 ttn* ,,1.2)
Y,;/'rr%i{
'94rn - Hyrolases '
cz// .
fu/hwn
S ,^ T4/, -PhYtoalexins
$
*
'(! ' -- vllu
O*idatire
cl.lvtr u u lJt -)7 lllu
bunst u u u i plant
induces cell
P l cl l l l ' utrl l death
"
, prllsVarwoffvts
_''
"-
"V/ :

3l Mechanism:
-The V wn,rfuftL \
. Plant
detects markers/elicitors frompathogen ) lnduces innate (basal)immunity Tt4aint'
5| "' S
t \ - Release ofROS, hydrolases, calose N
E\ - Whataresome examples ofelicitors? N
.- FlaglileilNi
Fragments ofueil
ul cellwalls, conserved
w4il5,uuiluervsu proteins
pruterns(shared
tsilareq oy majority
byaa rilajonly ofoac.,
or bac.)
A\
t\
. Pathogens $
$
e\ may produce virulence proteins to counteract innate immunity 1\
A.
\s
5,-\
t\ Delivered
-- llolirrorarl totha
fn inside
theincida offha
nf plant
thenlanl viahrna
rria typellllll secretion system--->
canralinn crrclam e.tu;t^,
---"> f,lnli/a4 +^ nnr)4ilA/)-f7btA
n SlWf#*
to\ ' Tocounteract virulence proteins, theplantproduces R-proteins* zfatlsalue/ rhln 'p:orrru {"
\ - Made tobindtovirulence proteins) induces hypersensitive response ( t/ \
\ . Release phytdabxins, ROS, RNS, Progammed celldeaih, Increased lignin . ..d, \t
b,/ z-sarurc -
oili a2 ;,-iinL
->rtal at py'z7<,ttsrutl/ be
aurt/ana firy;4it" i;-i'&;t;r']:'i'
lvcglilztrl. ,E-gpflrs -.,-.g.s
-Hypersensitive
response
onryinitiared .tU,
- Whathappenswhena plantis -^rl#,iini"KAffi#i"6k-W#"*fn'itti*4/u,/ns
notabletofendoffaninfectiontn* ,
theplant,
i'spreadsthroughout Systemicinfection vt4ftU?r 0( SP"af'tttuJ
in $ o e c'
t u
AGROBACTERIUM! A Slar'rr *lfr'!Y.,,,
SJyprr Pathoqenic? ,iget,letd \!ry
A.radiobacter ) noInfection wytory\t+
A,rizogenes ) Hairyroottumor ' r
A.Tumefaciens ) Undifferentiated
tumor/gall

TheBasic
-lnfection: Characteristics!
1)Plantsshouldbewounded toinfect
- Timeofinfection
iscrucial
too) optimum at12-18 hrsafterwounding
2)Temperature ) willnotinfect
sensitive athightemperatures
planttumor
3)Infected cellscangrowinmedia without hormones ) transformed
- REMEMBER: togrowuntransformed cells,needpresence ofhormones
inmedia
4)Tumor cellsproduce
opines
- Sugar andAmino Acidbonded together, releasedintothesoil
- OnlyAgrocanusethecompound asnutrients
go/ion'/'/""'
- Whatisneededforplantinfection?
) TiPlasm id -*fitlf d,s////,fut/vt {?
 4 Parts
of a Ti Plasmid:
- Framework
1)ORI= Origin
ofReplication
- Needed plasmid
toreplicate whenbacteria
divides
2)OpineCATABOLISIM genes
- Needed touseasa nutrition
Opine
toMetabolize source
3)VirGenes
- ProduceVirproteins tomobilize
needed andtransferT-DNA
. Makess COpy
OfT-DNA-, ,S i r-rir\)i.,*-u1A *" ,'
;
. MakeBridge syst6m)l; t '/:'
toplant(typelVsecretion
. Move
T-DNA toplantnucleus
' lnt'(futa&pefrLJ
/ ftfil fettoffJ e-

a CloserLook
4)TheT-DNA:

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

Fillinthechart:
Agrogenes Cvtokinin Auxin Growth
TmrTms- NoGrowth!
Tmr-Tms* + ntot *uue{'L
Tmr*Tms- {- .9't ot)irlgtl^A D (-

Tmr*Tms* (w.t.) + .|- uvt J tzu,ttt h aVA .t'1.)

dt [^^JL
- Canreproduce withUN-TRANSFORMED
results cellsbyadding the inghormones to media
(w.saltsandsucrose)
. Keytoregenerating/propagatirE
plantcellsintissuecultures
you
- Can describe thebasicsteps of plantpropagation? ,olnr/fto' a- /Ph'y'/lar
- Doyouplace incytokinin
thecallus orAuxin first?
| .+ Ailxrn Znd, 4+ok-rVtrv'r \s* I
PlantTransformation
A.Tumefacians
realize
Scientists canbeusedasa vectortotransform plantswithgenes oftheirchoosing!
- Needtoremove ryineandESPECIALLY oncogenesfrom T-DNA region . ^- ,. !.,
'tiffi"fr:l'tr;;T1u'ttm4'r7n;dtz
,*vflh'3'*K&*/';,trry*'f,itrFif
- T-DNAmodifications:
. Multiple Site(MCS)-
Cloning Restriction forgeneofchoice
sitesthatallowinsertion
. Selectable thetransformed
Todistinguish
marker- plantcells
genesforKanamycin
- Usually resistance (Kant) // -/,n ,"r4r4u/.,///r t.u/
4t'tt ;/"5
4hn,ts marl/f /'nf n7vA7fuatPt/ "' 'n'br*rrO/r; ,
i
1)WoundplantandapplyAgroculture cellstoagarplate[w/salts,
with 2)Apply
transformed carbonsource,
TiPlasmid
w/ geneofinterest hormones(aux& cyt),
andKanl.Allow todevelop
Callus

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

newplanttosoil
5)Transfer Process
Completed!Plantentirely
composed
ofcells
geneofinterest!
containing

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

. Because
T-DNA plasmid
issignificantly toworkwith!
easier
smaller,
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