Screening and cloning of the target genes immuno-regulated by glycyrrhizin using suppression subtractive hybridization technique / 解放军医学杂志

ABSTRACT

Objective To construct a subtractive cDNA library of genes differentially expressed in human lymphoma cell line Jurkat cells treated with glycyrrhizin (GL), and to clone genes associated with its immunological regulation, and to further elucidate the molecular immune mechanism of GL. Methods The mRNA was isolated from Jurkat cells treated with either GL or 0.9 percent sodium chloride as a control, then cDNA was synthesized. Suppression subtractive hybridization (SSH) method was employed to analyze the differentially expressed DNA sequence between the two groups. The cDNA was sequenced and analyzed in GenBank with Blast search after the amplification of the subtractive library by PCR. Results The amplified library contained 28 positive clones. Colony PCR analysis showed that there were 22 clones containing 200-1 000 bp inserts. Sequence analysis was performed, and the full length sequences were obtained with bioinformatics method. Altogether 11 kinds of encoding sequences were achieved, including interleukin-12, interleukin-18, and thymosin ?1, etc. Conclusions A subtractive cDNA library of genes differentially expressed in Jurkat cells treated with GL using SSH technique was constructed successfully, and it might give some new clues for the study of the immune regulation mechanism of GL.