类乌齐牦牛RNF216基因的克隆及组织表达分析

Cloning and Tissue Expression of RNF216 in Leiwuqi Yak

【目的】为获得类乌齐牦牛环指蛋白216抗体(ring finger protein 216,RNF216)基因的CDS区序列,并研究RNF216基因在类乌齐牦牛不同组织中的表达情况。【方法】试验以0.5岁龄类乌齐牦牛为试验材料,采用RT-PCR方法克隆牦牛RNF216基因CDS序列并进行生物信息学分析,利用荧光定量PCR(qPCR)方法检测RNF216基因在臀脂、臀肌、肝脏、肺脏、心脏和脾脏等组织器官中的表达情况。【结果】RNF216基因CDS区全长2610 bp,开放阅读框序列长2223 bp,共编码740个氨基酸。RNF216基因编码蛋白为不稳定亲水酸性蛋白,无信号肽和跨膜结构,主要分布在细胞核中。理化性质分析显示,牦牛RNF216蛋白存在24个丝氨酸、8个苏氨酸和9个酪氨酸磷酸化位点。一致性比较和进化树分析显示,牦牛与黄牛的遗传距离最近,与鼠类的遗传距离最远,且在进化过程中保守性强。组织表达分析显示,RNF216基因在类乌齐牦牛肺脏中的表达水平最高,其次是脾脏,然后是臀脂,在臀肌中的表达水平最低。【结论】本研究为进一步了解RNF216基因在调控牦牛肌肉生长发育方面提供参考数据。

【Objective】The sequence of CDS region of the gene encoding ring finger protein 216(RNF216)was obtained,and the expression of RNF216 gene in different tissues of Leiwuqi yak was studied.【Method】In this experiment,the 0.5-year-old Leiwuqi yak was used as the experimental material.The CDS sequence of yak RNF216 gene was cloned by RT-PCR and analyzed by bioinformatics.The expression of RNF216 gene in hip fat,gluteal,hepar,lung,heart and spleen was detected by real-time PCR(qPCR).【Result】The results showed that the CDS region of RNF216 gene was 2610 bp in length and the open reading frame sequence was 2223 bp in length,encoding a total of 740 amino acids.The protein encoded by RNF216 gene was an unstable hydrophilic acidic protein with no signal peptide and transmembrane structure,which is mainly distributed in the nucleus.Physical and chemical analysis showed that there were 24 serine,8 threonine and 9 tyrosine phosphorylation sites in the RNF216 protein.Consistency comparison and phylogenetic tree analysis showed that the genetic distance between yak and cattle was the closest,the genetic distance from the mouse was the farthest,and it was conservative during the evolution process.Tissue expression analysis showed that the expression level of RNF216 gene was the highest in the lungs of the Uzbek-like calves,followed by the spleen,which was the lowest in the gluteal muscle.【Conclusion】This study provides a reference for further understanding of the RNF216 gene in regulating the growth of yak muscle.