High Frequency Genetic Transformation System Of Medicago Truncatula Mediated By Agrobacterium Tumefaciens And Preliminary Study In Function Of MtRop6

Medicago truncatula is currently used as a model plant for investigating the symbiotic relationships between legumes and rhizobia. Genetic transformation is an indispensable technique to elucidate gene expression and regulation of plant growth and development. Establishment of high frequency transformation and regeneration system is important to discover novel functional genes from M. truncatula.The high embryogenesis line R108-1 has been used in present investigation. We have optimized different transformation factors and established an efficient Agrobacterium tumefaciens (GV3101) mediated genetic transformation system. In vivo transformation leaf explant through somatic embryogenesis developed intact seedlings. But rooting is still a bottleneck in the regeneration-transformation process of M. truncatula R108-1. We have used A. rhizogenes (MSU440) to mediate transformation for producing hair roots in sectioned regeneration seedlings. This step consumedly improved rooting efficiency from 5~7% to 40%. In addition, more than 60% regenerated seedlings can be induced to produce roots on M3 medium, and the produced root system has many lateral roots which contributes to survival of the seedlings in the pots.ROP GTPases have been implicated in regulating various plant cellular responses including defense against pathogens. Legume-rhizobium symbiosis, especially at the early stage in symbiosis, requires avoiding or suppressing the plant defense response. The MtRop6 gene was cloned from M. truncatula, and bioinformatics analyses showed MtRop6 possibly participated in ROS (reactive oxygen species) pathway in plant defense signal transduction. Our interest is investigating the function of MtRop6 during establishment of symbiotic interaction. So, we had constructed several clones to investigate the function of MtRop6 in plant defense and different symbiosis stages by using A. rhizogenes and A. tumefaciens transformed M. truncatula respectively. The preliminary result show that MtRop6 gene's expression reduced will cause the root hair development retardation, and the H2O2 accumulation obviously reduced in the shortened root hair.