基于RNA-seq技术的水杉半胱氨酸合成酶基因的克隆分析

Cloning and Analysis of Cysteine Synthase Gene Based on RNA-seq Technology in Metasequoia glyptostroboides

水杉(Metasequoia glyptostroboides)是世界著名的孑遗植物,但对其在分子水平研究的报道极少。为了获得与耐逆境胁迫有关的半胱氨酸合成酶基因,本研究以水杉原生种为实验材料,基于高通量转录组测序(high-throughput mRNA sequencing,RNA-seq)技术从全长转录组文库中克隆了一个半胱氨酸合成酶基因(MeGl-CSase),并对该基因作了进一步的生物信息学分析。结果表明,MeGl-CSase基因的全长cDNA为1702 bp,编码长度为465个氨基酸的半胱氨酸合成酶蛋白;该蛋白的分子量为49471.35 Da,总平均吸水值为-0.071,等电点为7.95;含有39个常见功能位点;二级结构包括11个折叠区、21个螺旋区以及一些卷曲区;三级结构预测结果进一步证明MeGl-CSase基因编码的产物是真核生物半胱氨酸合成酶蛋白。本研究结果可为水杉及其它物种的半胱氨酸合成酶蛋白基因的结构和功能研究提供参考。

Metasequoia glyptostroboides is a famous relic plant in the world,but only few studies at molecular level were reported.In order to obtain a cysteine synthase gene associated with stress tolerance,in this study,M.glyptostroboides was used as experimental material,and a cysteine synthase gene(named MeGl-CSase)was cloned from a full-length transcriptome library based on transcriptome sequencing(RNA-seq)technology.The cloned gene was further analyzed by bioinformatic analysis.The results showed that its full-length cDNA is 1702 bp and it encodes a cysteine synthase consisted of 465 amino acids;the molecular weight of the encoded protein is 49471.35 Da,grand average of hydropathicity is-0.071,and the isoelectric point is 7.95;there are 39 common functional sites;its secondary structure includes 11 folding regions,21 helical regions and some coil regions;the predicted tertiary structure further demonstrated that the product encoded by MeGl-CSase gene is a eukaryotic cysteine synthase protein.The results of this experiment could provide reference for the structure and function research of cysteine synthase protein genes in M.glyptostroboides and other species.