Reverse Genetics and Tilling Conclusions

Sensitive mutation discovery by TILLING is opening up new prospects for functional genomics in a wide array of crops, especially those for which sequence information is limited or those in which transformation is not available or efficient. Some of the plants in which TILLING has been reported include Arabidop-sis, lotus, wheat, maize, rice, barley, poplar, and soybean, with the list continuing to grow (Gilchrist and

Haughn 2005; Hohmann et al. 2005; Slade and Knauf 2005; Comai and Henikoff2006). Arcadia Biosciences has also successfully applied TILLING in tomato, lettuce, castor, peanut, soybean, maize and rice (Slade et al. unpublished). One reason for its increasing popularity across diverse plant species is the general applicability of chemical mutagenesis and lack of requirements for special constructs or transformation protocols. However, it can take considerable time to develop a mutagenesis treatment optimized for each crop, especially one in which lethality is balanced with an adequate mutation frequency for efficient throughput. In addition to identifying novel induced diversity, "ecoTILLING" can be used to discover naturally occurring SNPs and small indels in different plant varieties and landraces (Comai et al. 2004; Gilchrist et al. 2006). This is an especially attractive way to uncover diversity in plants that are less amenable to the usual TILLING strategy, such as those with long generation times or in crops that are propagated vege-tatively.

Thegenetic diversitythatiscreatedthroughchem-ical mutagenesis can range in effect from eliminating gene function entirely, to severely or slightly altering function, to having no effect at all. Such a range of potential phenotypes provides investigators with a full tool box for elucidating the function of unknown genes and can facilitate research of known genes where a reduction of function rather than a complete knock-out is desired. TILLING is also a non-transgenic method for introducing genetic diversity into domesticated varieties to provide novel alleles for commercially important traits in tomato and other Solanaceae and thus can be a powerful tool for crop improvement.

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