甜瓜CmGnT基因的克隆及非生物胁迫下的表达分析

Cloning and Expression Analyzing of CmGnT Gene in Melon Under Abiotic Stresses

为了探究非生物胁迫下甜瓜N–乙酰氨基葡萄糖基转移酶基因(CmGnT)的作用机制,根据甜瓜cDNA序列设计引物,利用RT-PCR技术克隆得到CmGnT。生物信息学分析表明,CmGnT的cDNA全长为2 193 bp,开放阅读框(ORF)为1 179 bp,编码392个氨基酸。CmGnT蛋白分子量约为46 kD,理论等电点(pI)为7.93。蛋白结构分析表明,CmGnT含有β–1,4–N–乙酰氨基葡萄糖转移酶结构域,属于糖基转移酶17家族。该基因编码的蛋白有1个强的跨膜螺旋结构。进化树分析表明,CmGnT氨基酸序列与黄瓜CsGnT(登录号:XP_004135275.1)亲缘关系最近,同源性为99.74%,与西瓜Cs Gn T的同源性为97.70%。实时荧光定量PCR分析结果表明,在NaCl、PEG、H_2O_2和ABA等非生物胁迫下的甜瓜叶片中CmGnT都显著上调表达,表明CmGnT受非生物胁迫诱导,推测其在甜瓜响09应非生物胁迫过程中起到重要作用。

The aim of this study was to explore the mechanisms of N-acetylglucosaminyltransferase gene （CmGnT） in melon under abiotic stresses. Primers were designed according to cDNA sequence of melon, and CmGnT was cloned by using RT-PCR technique. The full-length cDNA of CmGnT was 2 193 bp, and the open reading frame （ORF） was 1 179 bp, encoding 392 amino acids. The molecular weight of CmGnT protein was about 46 kD, and the theoretical isoelectric point （pI） was 7.93. Protein structure analysis showed that CmGnT contained the β-1,4-N-acetylglucosaminyltransferase domain, which belonged to the glycosyltransferases 17 family. The protein had 1 strong transmembrane helices. Phylogenetic analysis showed that CmGnT and CsGnT （accession number： XP_004135275.1 ） had the closest genetic relationship, and the homology was 99.74%. Homology with watermelon was 97.70%. qRT-PCR analysis showed that the expressions of CmGnT in leaf were significantly up-regulated under NaCl, PEG, H2O2 and ABA stresses, which indicated that CmGnTwas induced by abiotic stresses. The results suggested that this gene in melon played an important role in response to abiotic stresses.