Download Plant Transformation

Lecture 6
Marker genes and marker free
transgenic plants
Selectable Markers
• allow the selection of transformed cells, or
tissue explants
• by ability to grow in the presence of an
antibiotic or a herbicide.
• frequently used - kanamycin and
hygromycin
• used to follow the inheritance of a foreign
gene in a segregating population of plants
Selectable Markers
• Common selectable marker genes:
– NPTII - detoxifies kanamycin (antibiotic)
– bar - detoxifies glufosinate (herbicide)
– csr-1 - renders plants resistant to sulfonylurea
herbicides
– PMI - changes mannose to useable
carbohydrate
Screenable markers
• encode gene products whose enzyme activity can be easily
assayed
• allowing not only the detection of transformants
• also estimation of the levels of foreign gene expression in
transgenic tissue
• markers such as GUS, luciferase or -galactosidase allow
screening for enzyme activity by histochemical staining or
fluorimetric assay of individual cells
• can be used to study cell-specific as well as
developmentally regulated gene expression
Selectable markers
• marker genes linked to promoters
• Two main aspects
1.nucleic acid sequence
2.the gene product
• the coding sequence fused to promoters isolated
from T-DNA or the CaMV genome at the 5’ end,
and a polyadenylation signal
• able to exert stringent selection pressure on the
plant tissue
T-DNA based selectable marker excision system
LB
Ds
SM
Ds
Tp
LB&RB-Left and right border of T-DNA
Ds
Transposable elements
SM
- Selectable marker
Tp
- Transposase enzyme
G
- Foreign gene
G
RB
Marker free transgenics