Cloning And Expression Of S-adenosylmethionine Synthetase Gene From Saccharomyces Cerevisiae

S-adenosylmethionine(SAM) is an important metabolite in vivo, and takes part in more than forty kind of biochemical reactions, such as the gene expression, cell membrane fluidity and the synthesis of peptide. Clinical studies showed that SAM has curative effects on osteoarthritis, depression and the disorder of liver function. SAM has also been found to be an advanced health care product which can prevent cancer, cardiovascular diseases and anti-senility. There are many patients with hepatitis in our country, and the number of patients with depression or osteoarthritis is increasing along with the faster living rhythm, so SAM has a broader market. But we have not yet been able to industrial production in China. Though there are enormous SAM commodities to supply for the people needing, it can not be popularized because of its expensive price. Thus it is urge to explore a high producing and low-cost producing technology.The gene encoding sam2 was amplified from Saccharomyces cerevisiae 2B-41 chromosomal DNA using PCR technique on the base of the specific primers from Saccharomyces cerevisiae in GenBank. The PCR product was approximately 1200bp DNA segment. Using T-A cloning technique , the PCR product was cloned into the pUCm-T vector and the cloning plasmid pUCm-T-sam2 was constructed successfully. pUCm-T-sam2 and pET28a(+) were digested by EcoR I and Hind III double enzymes. The purified SAM2 gene was cloned into the expression vector pET28a(+), and the prokaryotic expression vector pET-sam2 was thus constructed. The recombinant plasmid pET-sam2 was transformed into competence E.coli BL21(DE₃) strain. The bacterium was induced by IPIG and its lysates were analyzed directly by SDS-PAGE. An approximate 47kD exogenous protein was observed on the...