摘要
[目的]对从江香猪TPM3基因进行扩增、克隆和序列分析。[方法]通过RT-PCR从猪背最长肌中扩增TPM3基因CDS区并进行克隆,利用DNAMAN和BioEdit等软件对克隆的序列进行分析。[结果]成功克隆了从江香猪TPM3基因并构建了pUCm-T-TPM3载体;获得了TPM3-1和TPM3-2两个序列,TPM3-1有两个碱基的差异,相似度达99.73%;TPM3-2有一个碱基的变化,同时插入了一段76 bp的片段,相似度达99.87%;密码子偏好性分析显示,UUG编码亮氨酸的频率为0.49%,而CUG编码亮氨酸的频率为2.89%,说明第49位碱基的转换可能提高蛋白的合成效率;RNA二级结构分析显示,碱基突变会影响TPM3基因RNA二级结构和最小自由能的变化;蛋白理化性质分析显示,氨基酸的改变对α螺旋、β折叠及转角等二级结构影响不明显。[结论]从江香猪TPM3基因的碱基突变可能影响原肌球蛋白的合成,为探究其对从江香猪肉质及种资源的开发利用提供试验依据。
[Objective]It was to clone and sequence analysis of Congjiang xiang pig TPM3 gene. [Methods]The CDS region of TPM3 gene in Congjiang xiang pig was cloned by RT-PCR and its sequence was analyzed using software such as DNAMAN and Bio Edit. [Results]The CDS region of TPM3 gene from Congjiang xiang pig was successfully cloned and the p UCm-T-TPM3 vector was constructed. We had also acquired two different sequences involving in TPM3-1 and TPM3-2. Sequence analysis results had showed that TPM3-1 had two different sites comparing original sequence and its similarity was 99. 73%.On the other sequence,TPM3-2 had a changed base and inserted a DNA fragment of 76 bp length as well as similarity is99. 87% comparing with the original sequence. The codon bias analysis showed that the UUG-encoded leucine frequency was0. 49%,while the CUG-encoded leucine frequency was 2. 89%,indicating that the 49 th base conversion may increase protein synthesis efficiency. RNA secondary structure analysis showed that base mutations affected the changes of TPM3 RNA secondary structure and minimal free energy. Analysis of protein physicochemical properties showed that amino acid changes had no significant effect on secondary structures such as α-helix,β-sheet,and turn. [Conclusion]The base mutation of TPG3 gene in Jiangxiang pig may affect the synthesis of tropomyosin. It provides experimental basis for exploring the development and utilization of Jiangjing pork meat and its resources.
作者
王赛楠
许厚强
赵佳福
李航
Sainan Wang;Houqiang Xu;Jiafu Zhao;Hang Li(Key Laboratory of Animal Genetics,Breeding and Reproduction in the Plateau Mountain Region,Ministry of Education;Key Laboratory of Animal Genetics,Breeding and Reproduction in Gnizhou Province;College of Life Sciences,Gnizhou University,Gniyang 550025,China)
出处
《生物技术》
CAS
2018年第4期307-312,共6页
Biotechnology
基金
国家科技支撑计划项目("贵州香猪特色性状评估及特色基因挖掘与功能研究"
No.2015BAD03B02-3)
贵州省黔科合重大专项("贵州畜禽种质资源保存创新与利用"
No.(2013)6008)
