Insertional mutagenesis: a Swiss Army knife for functional genomics of Medicago truncatula

https://doi.org/10.1016/j.tplants.2005.03.009Get rights and content

Legumes are second only to grasses in worldwide economic importance, and understanding their molecular genetics is vital to the breeding of important grain and forage legumes. Over the past decade, Medicago truncatula has been selected as a model plant in which to study biological processes that are unique and pertinent to legumes, and that cannot easily be studied in Arabidopsis. Here, we discuss the most common tools for introducing and analyzing genetic mutations in M. truncatula. Because transformation and regeneration are still bottlenecks in studying a legume species, large-scale insertional mutagenesis poses a major challenge in M. truncatula. We discuss the tobacco retrotransposon Tnt1 as a viable and attractive option for introducing multiple independent insertions per plant for saturation mutagenesis.

Section snippets

Post-genomic challenge

Since the first whole-genome sequencing report of a free-living organism, which was for Haemophilus influenzae in 1995 [1], we have witnessed exponential progression in whole-genome sequencing of prokaryotic and eukaryotic organisms, including flowering plants and humans. The explosion of sequencing data together with the development of excellent bioinformatics tools means that the availability of sequence information might no longer be a factor that limits our understanding of plant

Medicago truncatula as a model legume

Legumes are second only to grasses in their worldwide economic importance [2]. Apart from their direct use as an excellent source of dietary proteins for human consumption and their crucial importance as pastures and forage crops in animal and dairy production, legumes play a pivotal role in maintaining our ecosystem by fixing atmospheric nitrogen in symbiotic association with soil bacteria of the genus Rhizobium. This unique ability of legumes improves soil fertility by supplying a freely

Genetic tools for Medicago truncatula functional genomics

Developing efficient ways to create genetic mutations and the means to identify the mutated genes has been the fundamental approach to understanding gene function. Many different approaches are either currently available or being developed to generate M. truncatula mutants (Table 1). The two classical approaches that have been used successfully in Arabidopsis are mutating seeds with chemicals [using ethyl methane sulfonate (EMS)] and ionizing radiation (using fast neutrons, γ-rays and X-rays)

Concluding remarks

Tnt1 promises to be an ideal tool for large-scale insertional mutagenesis in M. truncatula. Tnt1-tagged M. truncatula lines serve multiple purposes, analogous to the Swiss Army knife – forward mutant screening when curated, reverse genetics screening of DNA pools and the establishment of a flanking sequence database for in silico gene cloning. However, it would be unrealistic to assume that Tnt1 tagging alone will be sufficient for saturation mutagenesis. The strengths of the various approaches

Acknowledgements

We thank Rick Dixon, Greg May and Choong-Min Ryu for critical reading of the manuscript, and Younsig Kwak for stimulating discussions. We also thank Rujin Chen and Giles Oldroyd for their helpful suggestions and for sharing unpublished work. This work was supported by The Samuel Roberts Noble Foundation.

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