Cloning And Function Analysis Of CkLEA4-1 And CkLEA6-1 From Caragana Korshinskii Kom.

In Inner Mongolia area,the common natural disasters of drought,chilling,salinity happen frequently.These calamities have a negative influence on the growth,development and yields of the local crop,also cause a great loss for the local agricultural economy.So the related studies of plant stress resistance become more and more important.LEA protein is a kind of functional protein expressed in plants under stress condition which attracts lots of researchers' attention in recent years.In this study,two LEA encoding genes which named CkLEA4-1 and CkLEA6-1 were cloned from Caragana korshinskii Kom.,and their biological function were analyzed.The concrete conclusions are as following:1.Two LEA encoding genes were cloned by RT-PCR method.The open reading frame?ORF?of CkLEA4-1 is 1119bp in length,encoded a protein of 373 amino acids;the ORF of CkLEA6-1 is 255bp,encoded a protein of 85 amino acids.Two LEA proteins were both hydrophilic proteins as revealed by DNAMAN software.Sequence and phylogenetic analysis indicated that CkLEA4-1 protein belongs to the Pfam LEA₄,without a conserved domain,and was named as CkLEA4-1;CkLEA6-1 contains a LEA₆ domain,therefore it was designated as CkLEA6-1.2.Real-time-quantitative PCR analysis showed that drought,dehydration and NaCl stress enhanced CkLEA4-1 expression at the transcriptional level,and CkLEA6-1 expression was mainly induced by cold stress.3.The germination rate of CkLEA4-1 overexpression Arabidopsis seeds had no difference with the WT under 400mM mannitol treatment,but the green cotyledon rate of the transgenic lines were higher than the WT.The root length of CkLEA4-1 overexpression Arabidopsis seedlings were significantly longer than the WT under 400mM mannitol and 150mM NaCl treatments,and the transgenic lines growth better than the WT seedlings under 400mM mannitol,and their fresh weight increase.The 4-week-old CkLEA4-1 overexpression Arbidopsis lines enhanced tolerance to drought compared with WT under drought treatment.The MDA content of the transgenic lines were significantly lower than that in WT.4.The root length of CkLEA6-1 overexpression Arabidopsis seedlings were significantly longer than the WT under cold stress treatment,and the seedlings fresh weight slightly higher than the wild type.The relatively conductivity rate of CkLEA6-1 overexpression Arabidopsis plasma membrane under cold stress treatment showed that there is no obvious difference between transgenic lines and WT under room temperature and 4?,but the relative conductivity of transgenic lines were slightly lower than WT at-20?.While the MDA content of CkLEA6-1 overexpression Arabidopsis was lower than that in WT after cold stress treatments.