Plant Transformation and Regeneration

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Plant Transformation and Regeneration

• Gene Delivery Methods
• Agrobacterium tumefaciens-mediated T-DNA
  transfer
• Biolistic (gene gun)-mediated gene transfer
• Expression Cassette Assembly
• Critical components for transgene expression
• Vector Choices
• Tissue Culture-Based Gene Transfer
• Agrobacterium-mediated soybean transformation
• Biolistic-mediated maize transformation
• Transgene Analysis Integration patterns

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Crown-Galls Tumors tumefaciens
(Courtesy from Sharon von Broembsen)
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(Courtesy from Sharon von Broembsen)
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T-DNA Transfer From Agrobacterium To Plant Cell
T-DNA
Wild Type
RB
LB
Plant cell
Chromosome
Ti-plasmid
T-DNA
Disarmed
RB
LB
Chromosome
Ti-plasmid
Binary vector
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VirA
VirA
VirA
VirA
Wounded sites
Bacterial chromosome
chvA chvB chvF
VirG
VirG
Ti-plasmid
VirD2
VirE
VirE
virB, virC, virD, virE1, virE2, virF, virH
Vir region
VirE
VirE
VirE
LB
RB
VirE
RB
LB
VirD1
Binary vector
VirE
VirE
VirE
VirE
B4
B10
B9
B1
VirD2
VirD2
B11
VirE
VirE
VirE
VirE
B6
B7
VirE
VirE
VirE
VirE
VirD2
VirE
VirE
VirE
VirE
Agrobacterium tumefaciens
Cytoplastic receptors
Plant Cell
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Biolistic PDS 1000/He
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Critical Components for Transgene Expression
MCS
RB
Binary vector
Expression Cassette
LB
T-DNA
GOI
Promoter
Terminator
Cloning vector
MCS
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Soybean Transformation and Regeneration Process
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Genomic Southern Blot of Transgenic Soybean
W-13-12
W-14-20
W-14-24
W-14-29
W-11-42
W-13-28
W-13-24
W-14-8
Kb 23.0 9.4 4.4
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Maize Transformation and Regeneration Process
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