藏红花酸糖基转移酶UGTCs4基因的克隆、生物信息学及表达分析  

Cloning, bioinformatics, expression mode analysis of crocetin glycosyltransferase UGTCs4

作　　者：赖成霞[1] 玛依拉 玉素音 高燕[1] 李春平[1] 刘忠山[1] 石必显[1] 王博[2] 王玮[2] LAI Cheng-xia;MayilaYusuyin;GAOYan;LI Chun-ping;LIU Zhong-shan;SHI Bi-xian;WANG Bo;WANG Wei(Institute of Economic Crops,Xinjiang Academy of Agricultural Sciences,Urumqi 830091,China;Institute of Applied Microorganism,Xinjiang Academy of Agricultural Sciences,Urumqi 830091,China)

机构地区：[1]新疆农业科学院经济作物研究所,新疆乌鲁木齐830091 [2]新疆农业科学院微生物应用研究所,新疆乌鲁木齐830091

出　　处：《中草药》2020年第4期1044-1051,共8页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金项目(C020409);新疆自治区高技术项目(201211107)。

摘　　要：目的以藏红花悬浮细胞为研究对象,克隆藏红花酸糖基转移酶(crocetin glycosyltransferase)UGTCs4基因,并对其进行生物信息学和表达分析。方法采用同源克隆法和5’RACE(rapid amplification of c DNA ends)技术克隆UGTCs4基因的全长c DNA序列。运用生物信息学方法对该基因进行分析,预测其编码蛋白的结构与功能,并通过半定量RT-PCR(semi-quantitative reverse transcription and polymerase Chain reaction)方法检测不同诱导物条件下的基因表达情况。结果UGTCs4基因的c DNA全长为1380 bp,编码459个氨基酸。分子进化树分析,该糖基转移酶基因与已知的藏红花酸糖基转移酶基因UGTCs2具有相同的糖基化藏红花酸的功能。RT-PCR的结果显示,UGTCs4基因能够响应吲哚乙酸(IAA)、脱落酸(ABA)、赤霉素(GA)、过氧化氢(H2O2)、茉莉酸甲酯(Me JA)多种处理,这些处理均能促使其进行转录。结论首次从藏红花悬浮细胞中分离并报道了UGTCs4的c DNA克隆,并证实UGTCs4对不同诱导子的响应情况,为进一步研究藏红花素的生物合成及表达调控研究奠定了基础。Objective With the aim to obtain crocetin glycosyltransferase UGTCs4 in cultured saffron suspension cell, and carry out its bioinformatics and expression mode analysis. Methods A homologous cloning strategy and 5’ RACE methods were adopted, the full-length cDNA sequence of a crocetin glycosyltransferase, designated UGTCs4(GeneBank number: KX398932), was obtained. The characteristics of physiochemical properties, structure and function of the deduced UGTCs4 protein were determined using a series of bioinformatics tools. Semi-quantitative PCR was used for gene expression analysis. Results The results showed that the full length c DNA of UGTCs4 was 1 380 bp in length and encoding a 459 amino acid;UGTCs4 had high identities(83.2%) with UGTCs2 protein from saffron;UTGTCs4 had the same evolutionary tree as UGTCs2. UGTCs4 transcripts were constitutively expressed in the leaves, stems, and roots. UGTCs4 gene could respond to multiple treatments of indoleacetic acid(IAA), abscisic acid(ABA), gibberellin(GA), hydrogen peroxide(H2 O2), and methyl jasmonate(MJA), which promoted its transcription. Conclusion cDNA of crocetin glycosyltransferase was cloned from suspension cells for the first time and the response of UGTCs4 to different inducers was confirmed. Molecular characterization of UGTCs4 will be useful for further functional determination of the gene involving in the crocin biosynthesis and expression regulation.

关 键 词：藏红花酸 糖基转移酶 克隆 表达分析 藏红花悬浮细胞 

分 类 号：R282.12[医药卫生—中药学]