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(P<0.01)。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。
To explore the diversity and functions of cuticular proteins in crustacean, full-length cDNA of cuticular protein-5 in freshwater prawn Macrobrachium nipponense （MnCP-5） was cloned using epidermis transcriptome by RACE technique. The bioinformatics, expression profile and effects of KK-42 for the ex- pression of MnCP-5 were analyzed in different molt stages. The full sequence is 796bp containing a com- plete 477 hp ORF which encodes 158 amino acids with RR motif, a calculated molecular mass of 16.46 ku and pI of 4. 22. Comparison of amino acid sequences in crustacean, MnCP-5 shared 41%--53% identity with others, and the maximal similarity of 76 % with Quantitative Real-time PCR （qRT-PCR） showed that swamp red prawn Procambarus clarkii Casp-2. The MnCP-5 was expressed on all 3 stages C, D02 and Da 4, while the relative expression level at stage Da-4 was 1400 times as the level at stage C with very signif- icant difference （P〈0.01）. KK-42 treatment can up-regulate the expression of MnCP-5 at stage Do 2, and the expression levels at 3, 6 and 48 h were 6.3, 4. 8 and 11.4 times as the levels in control group. KK-42 can importantly move forward the peak expression of MnCP-5 from stage Da-4 to D0-2, which is involved in one of the potential molecular mechanisms of KK-42 for shortening the molt cycle of juvenile freshwater prawn.
出处
《水产科学》
CAS
CSCD
北大核心
2017年第6期747-752,共6页
Fisheries Science
基金
河南省基础与前沿技术研究项目(142300410021)
关键词
日本沼虾
表皮蛋白
基因克隆
表达
KK-42
Macrobrachium nipponense
cuticular protein
gene cloning
expression
KK-42
作者简介
郑征帆（1990-），女，硕士研究生；研究方向：甲壳动物生长发育的体液调节．E-mail：zhengzhengfan@163．com．;宁黔冀（1964），女，教授，博士；研究方向：甲壳动物激素调控．E-mail：nqjnqjl964@163．com．
