Tadege, Million , Wen, Jiangqi , He, Ji , Tu, Haidi , Kwak, Younsig , Zhao, Patrick X. , Ratet, Pascal , Mysore, Kiran S. .
Large scale Tnt1 tagging in Medicago truncatula reveals preferential targeting of exons.
Legumes constitute one of the largest crop families and are second only to grasses in economic importance to mankind. Over the past decade, Medicago truncatula has been selected as one of two model legume species to study legume functional genomics. Retrotransposon tagging is a valuable tool for plants with large genomes as retrotransposition is replicative and results in multiple stable insertions per genome. We used the tobacco retrotransposon Tnt1 in M. truncatula to generate over 5,600 independent lines in tissue culture representing an estimated 140,000 insertion events. Tnt1 inserts on an average of 25 different locations per genome during tissue culture and insertions are stable during subsequent generations in soil. Analysis of over 2,000 Tnt1 flanking sequence tags (FSTs) showed that Tnt1 strongly prefers (78%) gene-rich regions. Within the euchromatic gene space, Tnt1 showed a 68.6% bias towards genes due to preference for coding exons, but it has neither target site specificity nor insertion hot spots. Forward genetics screening of 1,557 R1 lines resulted in identification of a wide range of visible mutant phenotypes in more than 28% of the regenerated lines with allelic preponderance. The strong preference of Tnt1 for exons and its ability to insert at multiple positions per genome suggests that approximately 12,172 lines are sufficient for a 90% gene tagging coverage in M. truncatula. This is in contrast to the 365,487 lines that would be needed to do the same using T-DNA tagging. Our data demonstrates that Tnt1 is a unique candidate for saturation mutagenesis in M. truncatula and other plant species with large genomes.
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1 - Samuel Roberts Noble Foundation, Plant Biology Division, 2510 Sam Noble Parkway, Ardmore, OK, 73402, USA
2 - Institut des Sciences du Vegetale, CNRS, Avenue de la Terrasse, 91198, Gif sur Yvette, France
Presentation Type: Plant Biology Abstract
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM