Cloning and sequence analyzing of squalene synthase gene from Eleutherococcus senticosus endogenetic Penicillium minioluteum P116-1a

Abstract:

[Objective] To clone squalene?synthase (SS) gene from Penicillium?minioluteum P116-1a isolated from Eleutherococcus senticosus. [Methods] Using rapid amplification of cDNA 5￠ ends (5￠ RACE) method, we isolated completed cDNA and DNA sequence of SS from P. minioluteum. The gene was analyzed and corresponding structure and functions were predicted by the bioinformatic method. Expression of SS was detected by RT-PCR and SDS-PAGE. [Results] The results showed that SS gene had 4 exons and 3 introns. The open reading frame was 1?416?bp encoding a protein of 471 amino acid residues. The predicted secondary structure composition for the protein contained about 67.73% α helixes, 5.31% extended strand, 2.97% β turns and 23.99% random coil. SS had conserved binding regions of squalene synthase and phytoene synthase and located in endoplasmic reticulum membrane. The SS amino acid sequence of P. minioluteum showed more than 90% homology with that of P. marneffei and Talaromyces stipitatus. Expression of P. minioluteum P116-1a SS gene varied in different tempreture. [Conclusion] The SS gene of P. minioluteum form E. senticosus was successfully cloned for the first time, providing a stable foundation for studying on mechanism of improving eleutheroside content by P. minioluteum P116-1a.

Key words:Penicillium minioluteum P116-1a, Squalene synthase, Clone, 5￠ RACE method