丹参SmCYP76S7基因的克隆、亚细胞定位和表达分析

目的 克隆获得丹参SmCYP76S7基因序列，并对其进行生物信息学和表达特性分析。方法 克隆获得SmCYP76S7 的cDNA及基因组DNA序列，运用生物信息学软件对该基因及其编码蛋白进行结构和理化性质分析。构建融合表达载体pEGAD-SmCYP76S7-eGFP，转化烟草后分析SmCYP76S7蛋白的亚细胞定位。利用qRT-PCR测定SmCYP76S7基因在各器官中的表达量，同时分别利用不同光质和茉莉酸甲酯（methyl jasmonate，MeJA）处理丹参幼苗和丹参毛状根，探究SmCYP76S7基因对光质和MeJA的响应。结果 SmCYP76S7基因包含2个外显子和1个1 506 bp的开放阅读框，编码501个氨基酸。该基因在根、茎、叶和花中均有表达，其中花中表达量最低。SmCYP76S7蛋白除定位于内质网外，还分布于多个细胞结构。SmCYP76S7基因的表达在红光处理和MeJA处理样品中显著上调，是典型的光和MeJA诱导基因。结论 SmCYP76S7基因是CYP76家族成员，结合其表达特性和同家族其他成员的催化活性，该蛋白很可能参与丹参酮类成分的生物合成，其具体的生物学作用值得进一步深入挖掘。

Cloning, subcellular localization and expression analysis of SmCYP76S7 gene from Salvia miltiorrhiza

Objective To clone the gene sequence of SmCYP76S7 and analyze its bioinformatics and expression characteristics. Methods The cDNA and genomic DNA sequences of SmCYP76S7 were cloned, and the structure and physicochemical properties of the gene and its encoded protein were analyzed by bioinformatics software. The fusion expression vector pEGAD-SmCYP76S7-eGFP was constructed, and the subcellular localization of SmCYP76S7 protein was analyzed after transformation into tobacco. The expression level of SmCYP76S7 gene in various organs was determined by qRT-PCR. Meanwhile, the seedlings and hairy roots of Salvia miltiorrhiza were treated with different light quality and (methyl jasmonate) MeJA respectively to investigate the responses of SmCYP76S7 gene to light quality and MeJA. Results The SmCYP76S7 gene contained two exons and one open reading frame of 1506 bp, encoding 501 amino acids. This gene was expressed in root, stem, leaf and flower, with the lowest expression level in flower. In addition to the endoplasmic reticulum, SmCYP76S7 protein was also distributed in several other cell structures. The expression of SmCYP76S7 gene was significantly up-regulated in red light and MeJA treated samples, it was a typical light and MeJA-inducing gene.Conclusion SmCYP76S7 gene is a member of the CYP76 family. Combined with its expression characteristics and catalytic activity of other members in this family, SmCYP76S7 probablely participates in the biosynthesis of tanshinones, and its detailed biological role is worth further investigation.