ABSTRACT
Objective: To clone Gynostemma pentaphyllum squalene synthase gene (GpSS1) and analyze its sequence and expression pattern as well as its regulation in response to MeJA. Methods: Primers were designed based on the sequence of GpSS (GenBank accession numbers: FJ906799) and GpSS1 was cloned by using RT-PCR method. Physicochemical properties and transmembrane regions of the deduced GpSS1 protein were predicted via Protparam and Tmpred programs, respectively. Conserved domains involved in catalytic activity of SS enzyme were identified using MotifScan and multiple sequence alignment was achieved using the BioEdit software. Expression pattern of GpSS1 and its regulation by MeJA were analyzed by quantitative real-time RT-PCR. Results: GpSS1 contains an open reading frame of 1 254 bp encoding a putative protein of 417 amino acids. The protein has an aspartate-rich motif and an endoplasmic reticulum membrane anchoring region in addition to three conserved domains involved in catalytic activity of SS enzyme. The expression level of GpSS1 in young leaves was the highest, followed by that in old leaves, and the lowest was in rhizomes. The expression of GpSS1 was significantly upregulated in G. pentaphyllum leaves sprayed with different concentration of MeJA. The greatest upregulation of GpSS1 occurred in G. pentaphyllum leaves treated with 50 μmol/L MeJA. In both young and old leaves, the transcription of GpSS1 gradually increased and then decreased to varying degrees at 6-96 h after MeJA treatment. However, the expression level of GpSS1 was always higher in young leaves than that in old leaves. Conclusion: The cloning of GpSS1 and analysis on the expression regulated by MeJA will be helpful for further elucidating the function of GpSS1 and its mechanism regulated by MeJA, as well as for improving the quality and content of gypenosides.