日本沼虾表皮蛋白-5基因全长cDNA克隆及表达分析

发布时间：2018-01-23 14:34

本文关键词： 日本沼虾表皮蛋白基因克隆表达 KK-　出处：《水产科学》2017年06期 　论文类型：期刊论文

【摘要】：为探究甲壳动物表皮蛋白多样性及其功能,根据表皮转录组资料,利用RACE技术扩增得到日本沼虾表皮蛋白-5基因的cDNA全长序列,分析其生物信息学、不同蜕皮时期的表达特征以及KK-42对其表达特征的影响。日本沼虾表皮蛋白-5基因的cDNA全长796bp,开放阅读框477bp,编码158个氨基酸,含RR基序,预测蛋白等电点为4.22,分子量为16.46ku;氨基酸序列比对发现日本沼虾表皮蛋白-5与克氏原螯虾Casp-2相似性最高,为76%,与其他甲壳动物表皮蛋白相似性为41%~53%。实时荧光定量PCR结果显示,日本沼虾表皮蛋白-5基因在3个时期——蜕皮间期、蜕皮前早期和蜕皮前晚期均有表达,其中在脱皮前晚期的相对表达量最高,为蜕皮间期的1400倍,与之有极显著差异(P0.01)。KK-42处理显著上调日本沼虾表皮蛋白-5基因在蜕皮前早期的表达,在处理后3、6h和48h表达量分别是对照组的6.3、4.8和11.4倍。KK-42诱导日本沼虾表皮蛋白-5基因的表达,使峰值前移,这可能是KK-42缩短日本沼虾幼虾蜕皮周期的机制之一。
[Abstract]:In order to explore the diversity and function of crustacean epidermal protein, the full-length cDNA sequence of the epidermal protein-5 gene of Macrobrachium japonicus was amplified by RACE based on the data of epidermal transcriptome. The bioinformatics, the expression characteristics of different molting stages and the effect of KK-42 on its expression were analyzed. The cDNA of the epidermal protein-5 gene of Macrobrachium japonicus was 796bp in length. Open reading frame 477bp, encoding 158 amino acids, containing RR motifs, predicted protein isoelectric point 4.22, molecular weight 16.46ku. Amino acid sequence alignment showed that the Casp-2 similarity between Macrobrachium nipponensis epidermis protein -5 and Prochelia claricarpa was the highest (76%). The similarity of epidermal protein with other crustaceans was 41 and 53. The results of real-time fluorescence quantitative PCR showed that the epidermal protein 5 gene of Macrobrachium japonicus was at three stages of ecdysis. The relative expression was highest in the early stage of premolting and in the late stage of molting, which was 1400 times of that in the interphase of molting. KK-42 significantly up-regulated the expression of epidermal protein-5 gene of Macrobrachium japonicus in the early stage of premolting. At 6h and 48h, the expression of epidermal protein-5 gene in Macrobrachium japonicus was induced by 6.3N 4.8 and 11.4 times as much as that of control group, and the peak value was moved forward. This may be one of the mechanisms by which KK-42 shortens the molting cycle of juvenile Macrobrachium japonicus.
【作者单位】：河南师范大学生命科学学院;
【基金】：河南省基础与前沿技术研究项目(142300410021)
【分类号】：S917.4
【正文快照】： 由于坚硬表皮的限制,甲壳动物必须经过蜕皮才能生长,蜕皮是一个周期性的动态过程,包括旧表皮降解和新表皮的形成。作为表皮组成成分的几丁质是和蛋白质以结合的形式存在,此蛋白即为表皮蛋白(CPs),是甲壳动物表皮中重要的结构性蛋白,由蜕皮激素的靶组织——表皮上皮细胞按照一

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