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Plant Transformation and Regeneration

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

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Plant Transformation and Regeneration

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  1. 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

  2. Crown-Galls Tumors tumefaciens (Courtesy from Sharon von Broembsen)

  3. (Courtesy from Sharon von Broembsen)

  4. 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

  5. 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 VirD1 LB Binary vector VirE VirE VirE VirE VirD2 B4 B10 B9 B1 VirD2 B11 VirE VirE VirE VirE B6 B7 VirE VirE VirE VirE VirD2 VirE VirE VirE VirE Agrobacterium tumefaciens Cytoplastic receptors Plant Cell

  6. Biolistic PDS 1000/He

  7. Critical Components for Transgene Expression MCS RB Expression Cassette Binary vector LB T-DNA Promoter GOI Terminator Cloning vector MCS

  8. Soybean Transformation and Regeneration Process

  9. Genomic Southern Blot of Transgenic Soybean W-13-12 W-13-24 W-13-28 W-14-20 W-14-24 W-14-29 W-11-42 W-14-8 Kb 23.0 9.4 4.4

  10. Maize Transformation and Regeneration Process

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