摘要
目的克隆版纳微型猪近交系aquaporin 3(AQP3)基因,并利用生物信息学方法分析其序列特征,研究其在猪各组织中的表达情况。方法从版纳微型猪近交系脾脏中提取总RNA,利用RT-PCR方法扩增猪AQP3编码区序列,将纯化的片段与pMD18-T载体连接,转化宿主菌DH 5α,筛选阳性克隆进行测序。并采用半定量RT-PCR方法分析版纳微型猪近交系不同组织中AQP3基因的表达情况。结果成功克隆出AQP3基因编码区全长873 bp的序列,GenBank登录号为HQ888860,其编码290个氨基酸。氨基酸序列同源性分析表明,与牛的AQP3基因同源性最大,达99%。多组织半定量RT-PCR研究表明AQP3 mRNA在BMI猪的脾脏、肾脏、肺、小肠中均有表达,其中脾脏中表达最高。结论成功克隆出了版纳微型猪近交系AQP3基因全长编码区,并对其编码的蛋白质功能进行了预测,为进一步的功能研究奠定了基础。
Objective To clone the Banna mini-pig inbred line（BMI） AQP3 gene and to predict its function by bioinformatics analysis and to obtain its tissue expression profile.Methods The total RNA of BMI spleen was extracted,and the AQP3 gene was amplified by RT-PCR.The purified PCR product was ligated with a pMD18-T and transferred into the bacteria DH5α for replication.Then some positive clones were identified by sequencing.The gene expression profile of different tissues in BMI was analyzed by semi-quantitative RT-PCR.Results The coding region sequence of AQP3 was correctly cloned（GenBank accession number HQ888860）.The encoding sequence is 873 bp and encodes a protein of 290 amino acids.The deduced amino acids sequence was aligned with the corresponding part of other animals and had the highest similarity 99% with cow.Semi-quantitative RT-PCR analysis of mRNA from different normal tissues indicted that AQP3 gene was expressed in the spleen,kidney,skin and lung of BMI and the expression level was higher in the spleen than in the other organs.Conclusions AQP3 gene has been successfully cloned from Banna mini-pig inbred line and its function has been predicted.These results will provide a useful foundation for further studies on AQP3 function.
出处
《中国实验动物学报》
CAS
CSCD
2011年第5期366-371,共6页
Acta Laboratorium Animalis Scientia Sinica
基金
国家自然科学基金项目(No.31160439)
关键词
版纳微型猪近交系
AQP3克隆
序列分析
组织表达
Banna mini-pig inbred line（BMI）
AQP3 gene cloning
Sequence analysis
Tissue expression
作者简介
王配（1987-），女，硕士研究生，主要从事分子生物学方面的研究。E-mail：peipei99999@126．com，Tel：15198779711。第一作者
霍金龙（1975-），男，实验师，博士研究生，主要从事动物分子遗传学方面的研究。E-mail：jinlonghuo@ynau．edu．cn，Tel：13708761403。第一作者
[通讯作者]曾养志，男，教授，主要从事动物遗传育种方面的研究，E-mail：bnwxzjjx@yahoo．com．cn
