Construction And Analysis Of Tomato BAC Library And EMS Mutants

Tomato (Solarium lycopersicum), as one of the most popular vegetable crops in China, is suffering serious diseases, including bacterial spot, tomato yellow leaf curl virus, downy mildew and so on. In order to facilitate identifying genes conferring diease resistance and important horticultural traits for developing elite varieties with disease resistance, the work described here focus on constructing a BAC library and developing mutants.The S. pimpinellifolium accession PI128216was used for genomic BAC library construction. High weight molecular DNA was isolated, digested and ligated into the vector PCC1BAC. A total of61,440clones were obtained. The quality of the library was examined by isolating plasmid from two thousand randomly selected clones and Pulse field gel electrophoresis. The results showed that the average size of DNA fragment was94kb. The rate of empty vector and recombinant was5%. Thus, the total size of DNA fragments in the library was about6,000Mb, which covered more than6X of the tomato genome. This library will be useful for positional cloning and chromosomal landing of functional genes.In order to enrich the resources for research materialsgenetic study, a mutant library was also constructed using the tomato accession PI128216. Two concentrations of ethylmethane sulfonate (EMS) were adapted for seed treatment based on preliminary experiments. A total of9,000seeds were treated with0.8*EMS, and15,000seeds were treated with1.0%EMS. Morphological variations were recorded in the M1and M2generations using the descriptors and data standard for tomato. Sixteen classes and33sub-classess including seed, plant size, plant growth habit, leaf morphology, leaf color, blooming stage, inflorescence, flower morphology, flower color, fruit size, fruit color, fruit amount, infertility, viviparity and resistance. A large amount of mutants were obtained and some plants showed multiple mutations. These mutants provide a source for genetic study.The M2plant of mutant were subjected to evaluation of resistance to bacterial spot race T3via infiltration inoculation. Unfortunately, none of the1,924mutants were susceptible to the pathogen, which might be due to the population size was not enough to discover the change of resistance.In conclusion, a BAC library and a mutant library were constructed using the tomato accession PI128216. These materials will provide bases for genetic study, map-based cloning, and breeding.