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Abstract Detail

Large Scale Technologies and Resources

Mathieu, Melanie [1], Huang, Shaoxing [2], Zhang, Zhanyuan [3], Somers, David [4], Clemente, Tom [5], Wang, Kan [6], Nguyen, Henry [7], Stacey, Gary [8].

T-DNA and Transposon Mutagenesis of the Soybean Genome.

Gene knockout by random, saturation mutagenesis of plant genomes with T-DNA or transposons is a widely used approach to identify genes and their functions. Our project focuses on transposon tagging of soybean genes using two transposon systems, the maize transposable element Ds and the tobacco retrotransposable element Tnt1. Twelve, activation-tagging Ds vectors were constructed, each containing various combinations of either the CaMV 35s promoter or individual seed-specific promoters (e.g., phaseolin, soybean seed lectin, or glycinin). Several hundred transgenic soybean lines harboring the different Tnt1 and Ds element constructs were generated by Agrobacterium mediated transformation. About 130 of these lines were subjected to molecular analysis by PCR walking or inverse PCR to identify T-DNA insertions. We obtained the PCR walking fragment for about 80% of these lines. Analysis of these transformants by Southern-Blot revealed that 50% of the lines were single copy lines, 30% were double copy and 20% carried more than 2 copies. The average T-DNA insertion number was 1.5 insertions per line. A GmGeneDB (Glycine max Gene Database) was developed to serve as a resource to the community. The long term goal of the project is the generation of sufficient transposon insertions so as to target any soybean gene. Among the mutant collection, we tagged a gene responsible for male fertility in soybean. This T-DNA induced mutation inhibits the formation of mature pollen preventing normal self-fertilization. The insertion mutation was localized in a gene encoding a protein similar to a putative strictosidine synthase, normally involved in indol-type alkaloid metabolism.
This project is funded by a grant from the United Soybean Board.

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1 - University of Missouri, Plant Sciences, Life Sciences Center Room 202, Columbia, Missouri, 65211, Usa
2 - University of Missouri, Dalton Cardiovascular Center
3 - University of Missouri, Transformation Core Facility
4 - Monsanto
5 - University of Nebraska-Lincoln, Dept. of Agronomy & Horticulture Plant Science Initiative/ Center for Biotechnology
6 - Iowa State University, Department of Agronomy
7 - University of Missouri, Department of Agronomy
8 - University of Missouri, Plant Sciences

reverse genetics
PCR walking
Flanking sequences
Mutant analysis

Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P42006
Abstract ID:573

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