运用生物信息学软件对湘油15 PGIP9基因的核苷酸、蛋白氨基酸序列进行分析,并对其蛋白结构进行预测.结果表明:湘油15 PGIP9编码区CDS长1011 bp,编码336个氨基酸的开放阅读框,分子量为37.5kDa,等电点为7.9.N端1～22个氨基酸是信号肽,且这一区域疏水性较强,具有5个潜在的N-糖基化位点.N端和C端还各具有4个参与二硫键形成的半胱氨酸残基.二级结构显示有11个α-螺旋,14个β-延伸和25个无规则卷曲.中心LRR结构域由6个串联的LRR基序组成.随后将PGIP9的CDS序列亚克隆到原核表达载体pET -32a(+)中,构建pET - 32a - PGIP9重组表达质粒,并转化E.coil BL21(DE3),25℃,终浓度为0.2 mmol/L和0.5 mmol/L的IPTG诱导2h,都成功的表达了融合蛋白pET - 32a - PGIP9,其分子量约为52kDa,发现主要以包涵体形式存在.没有可溶形式的蛋白表达.%Ribonucleotide and amino acid sequence of Xiangyou 15 PGIP were analyzed, the protein structure were predicted by biological information software. CDS of Xiangyou 15 PGIP9 code area was 1 011 bp, which encoded 336 amino acid of open reading frame with a molecular mass of 37. 5 kDa and isoelectric point of 7.9. 1 - 22 amino acids were predicted to the N - terminal signal peptide with stronger hydrophobic region. The deduced amino acids sequence contained 5 potential N - glycosylation sites and 4 cysteine residues on N - terminal and C - terminal respectively, which involved in constituting the disulfide linkages. Secondary structure displayed it contained 11 a - helices, 14 β -extension and 25 random coils. The center LRR structural domain was composed of 6 tandem LRRs motifs. Later, CDS of PGIP9 was subencoded into the prokaryotic expression vector pET - 32a ( + ) constructing recombinanl expression plasmid pET - 32a - PGIP9 and was transformed into Escherichia coli BL21 ( DE3) . The fusion protein pET - 32a - PG1P9 was successfully expressed under final concentration 0.2 mmol/L and 0.5 mmol/L IPTG at 25X1 inducing for 2 h. The molecular weight of fusion protein was 52 kDa, it mainly appeared as inclusion bodies, not appeared as soluble protein.
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