Cloning and expression of cysteine synthase gene from <I>Po-lygonum sibiricum</I> Laxm.

Abstract

Cysteine synthase is a key enzyme for restricting plant cysteine synthesis. Cysteine synthase gene, designated PcCSase1 (GenBank accession no. EU597481), was successfully isolated from Polygonum sibiricum Laxm. by RACE technique. This gene was 1 260 bp in full-length and encoded a peptide of 382 amino acids. Based on bioinformatic analysis, PcCSase1 is a cytoplasm cysteine synthesis and contains a 16 amino-terminal (N-terminal) signal peptide, which led the PcCSase1 to go to the cytoplasm. The results obtained through homologous sequence analysis indicated that PcCSase1 mature protein was highly conserved in plants, which shared approximate 90% in the amino acid sequence. Expression analysis by RT-PCR showed that PcCSase1 gene was expressed in leaf, stem and root with the largest expression in leaf. Under 3% NaHCO3 stress, the largest expression of PcCSase1 gene was detected in leaf, stem and root at the second day following stress. PcCSase1 gene was inserted into pYES2 and transformed into yeast cells (Saccharomyces cerevisiae). The contents of the glutathione in the recombinant yeast and the cysteine in the medium were increased. INVSc1-pYES2-PcCSase1 was more tolerant to salt treatment than INVSc1-pYES2 and the former survival rate was higher than that of the later under the stress of 10% NaHCO3 and 5 mol/L NaCl. These results proved that PcCSase1 gene may confer high salt-tolerance.