摘要
为了克隆TNNI基因家族,预测其蛋白结构和功能,并分析其在牦牛不同组织中的表达差异,以0.5岁健康的类乌齐牦牛为试验材料,采用RT-PCR技术克隆牦牛肌钙蛋白Ⅰ基因家族的CDS区序列并进行生物信息学分析,利用实时荧光定量PCR技术检测TNNI基因家族各成员的mRNA表达水平。结果表明,TNNI1、TNNI2和TNNI3基因CDS区大小分别为564,549,639 bp,分别编码187,182,212个氨基酸残基。预测分析结果显示:TNNI基因家族编码的蛋白质均为偏碱性蛋白,蛋白结构不稳定,均无跨膜结构域,无信号肽,属非分泌型蛋白;二、三级结构均以α-螺旋为主,均含有肌蛋白超家族保守结构域。系统进化树分析表明:类乌齐牦牛TNNI1基因与水牛的亲缘关系最近,其次是绵羊、黄牛,与小鼠亲缘关系最远;TNNI2和TNNI3均与黄牛、水牛的亲缘关系较近,与其他亲缘关系较远。实时荧光定量PCR结果显示,TNNI1和TNNI2基因在臀肌中的相对表达量最高,且TNNI1基因在心脏、肝脏和肺脏中的表达量显著高于TNNI2基因(P<0.05),TNNI3基因在心脏中表达量较高,而在肺脏、臀肌和肝脏中表达量低。
In order to clone the TNNI gene family,predict its protein structure and function,and analyze its expression differences in different yak tissues,this study used 0.5-year-old healthy Leiwuqi yak as the experimental material,and the CDS of the yak troponin I gene family was cloned by RT-PCR and then the sequences were analyzed using bioinformatics,the mRNA expression level of TNNI family genes were determined using Real-time PCR.The results showed that the CDS of TNNI1,TNNI2 and TNNI3 were 564,549 and 639 bp,encoding 187,182 and 212 amino acid,respectively.The results of predictive analysis showed that the proteins encoded by the TNNI gene family were all alkaline proteins,the protein structure was unstable,no transmembrane domain,no signal peptide,and non-secretory protein;the second and third-order structures wereα-helix mainly,both contained the Troponin superfamily conserved domain.Phylogenetic tree analysis showed that the Leiwuqi yak TNNI1 gene had the closest associated with that of Bubalus bubalis,followed by Ovis aries and Bos taurus,and the furthest relationship with Mus musculus;TNNI2 and TNNI3 were closely related to Bos taurus and Bubalus bubalis,and distant with the others.The Real-time quantitative PCR results showed that the relative expression of TNNI1 and TNNI2 genes were the highest in the gluteal muscles,and the expression of TNNI1 in the heart,liver and lung was significantly higher than that of TNNI2(P<0.05),the TNNI3 gene was expressed in the heart at a higher level,but the expression level was low in the lung,gluteal muscle and liver.
作者
向娅
柴志欣
王吉坤
信金伟
王会
王嘉博
武志娟
钟金城
姬秋梅
XIANG Ya;CHAI Zhixin;WANG Jikun;XIN Jinwei;WANG Hui;WANG Jiabo;WU Zhijuan;ZHONG Jincheng;JI Qiumei(Key Laboratory of Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization,Sichuan Province and Ministry of Education,Southwest Minzu University,Chengdu 610041,China;State Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement,Lhasa 850000,China)
出处
《华北农学报》
CSCD
北大核心
2020年第2期228-238,共11页
Acta Agriculturae Boreali-Sinica
基金
西南民族大学研究生创新型科研项目(CX2019SZ72)
国家肉牛牦牛产业技术体系项目(CARS-37)
西藏自治区财政专项项目(XZNKY-2019-C-052)
青藏高原生态畜牧业协同创新中心开放基金项目(QZGYXT02)。
