Cloning and sequence analyses of Eleutherococcus senticosus GAPDH / 中草药

ABSTRACT

Objective In order to make the gene a valuable internal control gene, GAPDH of Eleutherococcus senticosus was cloned and analyzed in sequence. Methods Total RNA of E. senticosus was extracted using improved isothiocyanate method. Part sequence of GAPDH was coined by RT-PCR, and the sequence was acted as internal control gene for semiquantitative PCR. Results Length 627 bp of E. senticosus GAPDH was cloned, speculating coding 209 amino acids. To compare the amino acid sequence of £. senticosus GAPDH with those of Panax notoginseng, P. ginseng, and Arabidopsis thaliana, the amino acid homology was 97%, 93%, and 93%, respectively, and the nucleotide homology was 94%, 86%, and 84%, respectively. When the sequence acted as internal control gene, the semiquantative PCR has benign amplification effect and good reproducibility. Conclusion The cDNA clone of E. senticosus GAPDH is first reported, the results prove that the sequence is able to be internal control gene for analysis on gene expression. This study could make a foundation for the key enzyme expression and regulate mechanism analysis in eleutheroside biosynthesis pathway.