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Genetic Transformation of Plants:
Methods and Approaches to
Develop Transgenic plants
Steps in producing Transgenic
-A source of foreign DNA containing the desired gene
-A vector that carries the gene
-Means to introduce the vector into host plant
Methods to create Transgenics
- Agrobacterium mediated transformation
- Electroporation
- Biolistic methods for direct entry
- Protoplast fusion
Agrobacterium mediated Transgenic
ANIMATIONS REMOVED
Steps in T-DNA integration
Plant wound --> phenolics --> sense
by VirA--> signal passed to VirG -->
VirG activates
expression of other vir genes --> the
products of vir genes help to excise TDNA from the Ti-plamsid
Identify the right statement: during Agrobacterium mediated transformation
1.
2.
3.
4.
T-DNA from bacterial genome gets integrated into the host genome
VirG gene from host genome gets integrated into the bacterial genome
T-DNA from host genome gets integrated into the bacterial genome
VirG from bacteria gets integrated into the host genome
Identify the correct sequence of steps in T-DNA integration
1. Plant wound phenolics  sense by VirA signal passed to VirG  T-DNA excise
2. Phenolics  plant wound  sense by VirA signal passed to VirG  T-DNA excise
3. Plant wound  phenolics  sense by VirG  signal passed to VirA  T-DNA excise
4. Plant wound  Signal passed to VirG phenolics  sense by VirA T-DNA excise
Identify the correct statement: For gene gun method of transformation
1.
2.
3.
4.
you need bacteria with Ti-Plasmid
A simple plasmid DNA with expression promoter can be used
A plasmid with T-DNA in needed for host genome integration
No –TDNA is required
Identify the correct statement: The agrobacterium based plasmid should
Maintain
1.
2.
3.
4.
Two antibiotic resistance genes: one for agro selection and for plant selection
Three antibiotic resistance genes: one for agro, one for E. coli, and another for plant
One antibiotic resistance gene: one for agro selection
Two antibiotic resistance gene: one for E. coli selection and another for plant selection
Identify the correct statement: Protoplast refers to
1.
2.
3.
4.
Plant cells resulting from plasma membrane removal
Plant cells resulting from enzymatic breakdown of protoplasm
Plant cells resulting from enzymatic breakdown of cell wall
Plant cells resulting from the enlarged vacuole
Identify correct statement: during Plasmid DNA isolation, EDTA is added
In the P1 buffer
1.
2.
3.
4.
To inactivate RNAse
To inactivate DNAse
To inactivate proteases
To inactivate cellulases
During Plasmid DNA isolation, the separation of bacterial chromosomal
And plasmid DNA occurs as
1.
2.
3.
4.
Chromosomal DNA stays in the supernatant, the plasmid DNA precipitates
Both DNA stay in the supernatant
Chromosomal DNA precipitates while the plasmid DNA stays in the supernatan
Both DNA precipitates with different affinities
Identify the correct statement: During PCR of NUD7 gene,
the forward and reverse primers represent
1. The start of and the end of the gene sequence respectively
2. The end of and the start of the gene sequences respectively
3. The start of and the End of the plasmid DNA respectively
4. The middle section of and the start of the NUD7 gene sequence respectively