1. Ti plasmid derived vector system
Lecture 3
Ti plasmid derived vector system

2. Ti Plasmid T-DNA region DNA between L and R borders is
transferred to plant
as ssDNA;
T-DNA encoded genes can be substituted by target genes
Tumor-
producing
genes
Opine catabolism
Virulence region
ORI

6. Ti-plasmid based vectors
Binary systems
Co-integrated vectors
Needs 3 vectors
Needs 2 vectors:
Disarmed Ti plasmid
capable for infection
Disarmed Ti plasmid
with gene of interest
(no vir genes)
Helper vector
for infection
(with vir genes)
Form
co-integrated plasmid
after homologous
recombination on T-DNA
Intermediate vector
with T-region
and gene of interest
(transferred by conjugation)
Helper vector
for transfer of
intermediate plasmid into A.tum

9. Co-integrated vectors (hybrid ti-plasmids)
Right now
rarely used
DISADVANTAGES:
1) Long homologies required between the Ti plasmid
and the E. coli plasmids (pBR322 based Intermediate vectors)
making them difficult to engineer and use
2) Relatively inefficient gene transfer compared to the binary vecto

12. Ti plasmid vector systems are often working as binary vectors
T DNA region removed
Gene of interest
Plant selectable marker
HELPER
plasmid
Disarmed Ti
plasmid
Virulence
region
Bacterial selectable
marker
ori for A. tum
ori for A. tumefaciens
ori for E.coli
DISADVANTAGE: Depending on the orientation,
plasmids with two different origins of replication may be unstable in E. coli
ADVANTAGE: small vectors are used, which increases transfer efficiency
from E. coli to Agrobacterium.
No intermolecular recombination is needed