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作者	单位
方建，牟利，李鹏，张鹍飞，刘新春，袁金娥，冯宗云	（四川农业大学农学院植物遗传育种学系大麦青稞研究中心，四川成都 611130）

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中文摘要:

为进一步研究查尔酮合酶（Chalcone synthase，CHS）在黄酮化合物代谢过程中的作用，以青稞94-19-1为材料，通过同源克隆技术分离CHS基因，并对分离得到的CHS基因进行生物信息学分析及原核表达。结果表明，从青稞94-19-1中克隆得到的CHS基因编码区长为1 197 bp，编码398个氨基酸。生物信息学分析表明，该基因编码的蛋白质分子量为43.479 kDa，预测等电点(pI)为5.92，是酸性蛋白；该酶蛋白体外红细胞中的半衰期为30 h，不稳定系数(II)大于40，属于不稳定蛋白；平均亲水指数为-0.090，是亲水性的蛋白。SDS-PAGE检测结果表明，将该基因克隆到表达载体pET-32a上，并在大肠杆菌BL21中表达，可得到64 kDa左右的融合蛋白，在IPTG诱导浓度为1.0 mmol·L^-1时，最佳诱导时间为3 h。

英文摘要:

In order to research the role of chalcone synthase gene in flavonoids metabolic pathway,Qingke “94-19-1” was used as the material for homologous cloning in this study to clone CHS gene which was then studied by bioinformatics analysis and prokaryotic expression.A sequence of open reading frame (ORF) with 1 197 bp encoding 398 amino acids was obtained.Sequence analysis showed that the emzyme HvCHS was an unstable,hydrophobic and acid protein with the molecular weight of 43.479 kDa,instable coefficient of 40.10(>40),the half-life of 30 h,the average hydrophilic coefficient of -0.090 and theoretical pI of 5.92.Prokaryotic expression of pET-32a-HvCHS,which was formed from the expressive vector pET-32a linked with the encoding sequence of HvCHS gene,was carried out in E.coli BL21.The fusion protein with 64 kDa was expressed including the protein encoded by the gene HvCHS and the tag protein of expressive vector pET-32a.The optimal conditions for prokaryotic expression were 1.0 mmol·L^-1 for IPTG concentration and 3 h for expression time.

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