Cloning And Functin Identificatin Of MrSKIP Genes In Medicago Ruthenica L.

In this researches Medicago ruthenica seedlings was treated with MS liquid nutrient medium that contained 20%PEG 6000 to simulate the environment of salt stress.Total RNA of the Medicago ruthenica seedlings which had been treated 6 hours was selected as a template for reverse transcription,and the full-length cDNA stress related pre-mRNA splincing factor gene MrSKIP was cloned by using the RACE technique and RT-PCR method.Analysis of the sequence is conducted and the results show that the full length of MrSKIP cDNA is 2280bp.It contains an open reading frame of 1833bp,encodes 610 amino acids,with a S-N-W-K-N polypeptide,and has a molecular weight of 68.86145 KDa and an isoelectric point of 8.95.The typical domain SNW_SKIP was confirmed according to bioinformatic analysis.Furthermore,MrSKIP gene was cloned from medicago ruthenica genome DNA,and sequence analysis showsed no introns in it.We interfered MrSKIP expression at the roots of Madicago truncatula(108 R)by means of tasiRNA technology and Agrobacterium rhizogenes transformation.Drought,salt and low-temperature stress was then respectively applied to the roots of the transgeneic plants to study the gene function of MrSKIP.Defects of length and fresh weight of the roots were detected compared with the control.It can be preliminarily concluded that the decrease of MrSKIP expression influenced the resistance of drought,salt and low-temperature stress,with draught stress resistance weakened greatly.