Cloning And Function Analysis Of PEAMT Gene From Suaeda Liaotungensis K.

Phosphoethanolamine N-methyltransferase(PEAMT) catalyzes methylation of phosphoethanolamine(P-EA), producing phosphocholine(P-Cho) which is dephosphorylated to free choline. Glycine betaine (GB) is synthesized by the two-step oxidation of choline. With respect to protection against osmotic stress, GB is regarded as being a kind of small, non-toxic osmoprotectants. GB acts as an osmoprotectant by maintaining the cell turgor and driving gradient for water uptake under stresses. GB can stabilize both the quaternary structure and function of enzyme, complex protein such as the photosystemⅡ(PSⅡ)and other macro-molecules and protect the highly ordered membrane structure.The main factor limiting GB accumulation in plants is the endogenous choline supply. After PEAMT gene is cloned and then transferred into plants via genetic engineering, the content of choline and glycine betaine could therefore be enhanced which plays an important role in improving salt tolerance.In the present study, PEAMT cDNA had been cloned from Suaeda liaotungensis, transferred into wild type tobacoos and the content of choline and betaine in PEAMT transgenics plants were tested. The main results were as follows:1. The full length of PEAMT cDNA was obtained through RT-PCR and RACE approaches. PEAMT cDNA is 1688bp, including a 34 bp 5′UTR, a 169 bp 3′UTR and a 1485 bp ORF encoding a 495-amino-acid polypeptide which was 99.6%, 84.82%,85.43% and 84.29% to PEAMT sequences of Suaeda japonica,Beta vulgaris,Spinacia oleracea,Atriplex nummularia in amino acid homology respectively.2. The PEAMT ORF was obtained and the plant expression cassette pBI121-PEAMT was yielded. The PEAMT construct was mobilized into Agrobacterium tumefaciens strain LBA4404 by the liquid nitrogen freeze thaw method for plant transformation.3. The PEAMT gene was transferred into wild type tobaccos (Nictiana tabacum L. cv. 89) via Agrobacterium mediation. PCR analysis showed that PEAMT transgenic tobacoos were obtained. Choline and betaine content were determined by Chemistry Colorimetry which were higher than that in wild type tobacoos. Overexpressing PEAMT from Suaeda liaotungensis increased the choline content in transgenic plants and moreover improved the betaine content in those.