Functional Analysis Of MsYSL6 And MsYSL7 Genes In Medicago Sativa

The Yellow strip-like protein（YSL）family has important and extensive functions.Studies on YSL protein in Alfalfa are few and the mechanism of action is still unclear.The establishment of alfalfa regeneration system is not perfect.In this study,homozygous screening,submicroscopic structure observation and photosynthetic parameter determination were carried out on MsYSL6 and MsYSL7 transgenic tobacco.The response of transgenic tobacco to abiotic stress and metal stress was analyzed;the effects of 2,4-D and 6-BA on callus formation in Medicago sativa Zhaodong were studied.The main research results are as follows:1.Screening and analysis on offspring of MsYSL6 or MsYSL7 transgenic tobaccoThe seeds of MsYSL6 or MsYSL7 transgenic tobacco were generational screened by Km medium and T₁ of T₂ and T₃ generation resistant transgenic lines were obtained;Seven MsYSL6 transgenic T2 lines and seven MsYSL7 transgenic T3 lines were identified by PCR.Compared with WT,the number of thylakoids and starch grains in the chloroplasts,chloroplast content,and photosynthetic parameters in the transgenic tobacco of Ms YSL6 or MsYSL7 increased increased.The photosynthetic capacity of transgenic tobacco was improved by tranformation of MsYSL6 or MsYSL7.2.Response of MsYSL6 transgenic tobacco to abiotic stressThe yellowing of MsYSL6 transgenic tobacco leaves was lower than WT under the stress of salt,alkali;the seed germination rate and root length of MsYSL6 transgenic tobacco were lower than WT under drought stress.Tranformation of MsYSL6 improved the tolerance in tobacco to salt,alkali,drought stress.3.Response of MsYSL6 and MsYSL7 transgenic tobacco to metal stressThe yellowing of Ms YSL6 and MsYSL7 transgenic tobacco leaves was lower than WT under the stress of copper or cadmium;the germination rate,growth status,root length,fresh weight and chlorophyll content of MsYSL6 and MsYSL7 transgenic tobacco was significantly higher than thatof WT under copper or cadmium stress;Small degree of oxidative damage.Tranformation of MsYSL6 and MsYSL7 improved the tolerance in tobacco to copper or cadmium stress.4.Establishment of tissue culture system of Medicago sativa ZhaodongUsing 2,4-D and cytokinin 6-BA as induction hormones,callus induction was carried out with cotyledon nodes,leaves and stems of zhaodong alfalfa as explants.It was found that the cotyledon node of Medicago sativa Zhaodong was the best explant.MS+2.0 mg/L 2,4-D+0.5 mg/L 6-BA was the optimal induction medium,and the cotyledon node induction rate reached the highest value.The callus was differentiated on MS+0.5 mg/L 6-BA+30 mg/L sucrose medium,and 1/2 MS+15 mg/L was the optimal rooting medium.Regeneration plants of Medicago sativa Zhaodong was obtained,and a stable and efficient regeneration system of Medicago sativa Zhaodong was established.