大蒜半胱氨酸合成酶的mRNA表达及生物信息学分析

doi:10.13560/j.cnki.biotech.bull.1985.2016.08.015

摘要/Abstract

摘要： 半胱氨酸合成酶的产物半胱氨酸是植物中含硫氨基酸的重要来源,大蒜中含硫氨基酸丰富,研究大蒜半胱氨酸合成酶的性质及生物学功能以明确其在大蒜硫代谢中的作用。利用生物信息学技术分析从NCBI数据库中获得的3个大蒜的半胱氨酸合成酶基因（AsGCS2、AsGCS3和AsGCS4）的开放阅读框,预测其蛋白序列、分子量大小、蛋白质特性、亚细胞定位、系统进化等特征;通过荧光定量PCR分析了3个半胱氨酸合成酶的组织表达特性。结果显示,AsGCS2、AsGCS3和AsGCS4的开放态读码框长度分别为1 152 bp、1 296 bp和1 236 bp;其编码蛋白的理论相对分子量分别为40.6 kD、34.1 kD和36.1 kD。AsGCS2被亚细胞定位于叶绿体中,而AsGCS3和AsGCS4均被定位于细胞质中。氨基酸比对结果表明,AsGCS2与AsGCS3相似度为70%,与AsGCS4相似度为59%;而AsGCS3与AsGCS4相似度为68%。进化分析图表明,AsGCS2属于Bsas2亚家族,AsGCS3属于Bsas1亚家族,AsGCS4属于Bsas6亚家族。荧光定量PCR结果表明,大蒜不同CSase的组织特异性是不同的,AsGCS2在叶中表达量最高,ASGCS3在根中表达量最高,而ASGCS4在叶和根中都有较高的表达量。3个半胱氨酸合成酶基因分别属于不同的Bsas亚家族,它们的组织表达特性也不相同,它们在大蒜不同组织中的半胱氨酸合成途径中起作用。

关键词: 大蒜, 半胱氨酸合成酶, 荧光定量PCR

Abstract: The cysteine synthesized by cysteine synthase is an important source of amino acids containing sulfur in plants. The amino acid with sulfur in garlic is abundant,thus we studied the properties and biological functions of garlic cysteine synthase in order to clarify its role in the metabolism of garlic. Three cysteine synthase genes of garlic,AsGCS2,AsGCS3 and AsGCS4were found from NCBI database,and then bioinformatics analysis and RT-PCR were conducted. The lengths of their open reading frames were 1 152 bp,1 296 bp,and 1 236 bp,respectively. The theoretical relative molecular weights of their encoded enzymes were 40.6 kD,34.1 kD,and 36.1 kD,respectively. By subcellular localization,AsGCS2 was localized in the chloroplasts,while AsGCS3 and AsGCS4 were localized in the cytoplasm. The results of the amino acid sequence alignment revealed that the similarity of AsGCS2 with AsGCS3 was 70%,and that with AsGCS4 was 59%,while it was 68% between AsGCS3 and AsGCS4. Phylogenetic analysis showed that AsGCS2 belonged to Bsas2,AsGCS3 belonged to Bsas1,but AsGCS4 was different from all known cysteine synthases,possibly belonging to Bsas6. RT-PCR expression analysis indicated that the expression levels of CSase varied in various tissues. The expression level of AsGCS2 was the highest in the leaves,the highest expression of AsGCS3 was in the root,and AsGCS4 expressed highly in both leaves and roots. In conclusion,3 cysteine synthase gene AsGCS2,AsGCS3,and AsGCS4 belong to the different subfamilies of Bsas,their tissue expression characteristics were not the same,and they play a role in different tissues of garlic in cysteine biosynthesis.

Key words: Allium sativum, cysteine synthase, qRT-PCR

郭天璐, 张欢欢, 杜建中, 郝曜山, 王亦学, 孙毅. 大蒜半胱氨酸合成酶的mRNA表达及生物信息学分析[J]. 生物技术通报, 2016, 32(8): 96-102.

GUO Tian-lu, ZHANG Huan-huan, DU Jian-zhong, HAO Yao-shan, WANG Yi-xue, SUN Yi. mRNA Expression and Bioinformatics Analysis of Cysteine Synthase in Allium sativum[J]. Biotechnology Bulletin, 2016, 32(8): 96-102.

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