Dai Jun-Yong, Cao Xiang-Rong, Shi Lei, Zhang Xi-Ran, Xu Chun-Mao, Hu Jun
College of Life Science, Nanjing Normal University, Nanjing 210097, China.
Yi Chuan. 2005 Jan;27(1):95-100.
In this study, partial fragments of potassium ion channel gene were amplified using the genomic DNA of muntjak, reevesi, crinifrons, and Elaphodus cephalophus. The PCR products were ligated to the plasmid of pMD18-T Vector by the method of direct T-A cloning. The positive clones were identified by colony PCR. The sequences of the recombinant clones were determined using M13-47/RV-M universal primers and aligned by the software CLUSTALW. The nucleotide divergences of exon were 0.90%-1.44% among three species of Muntiacus, 0.90%-1.26% between E. cephalophus and each of Muntiacus deer. In the nucleotide of intron there is 0%-1.22% difference among these muntjac deers, and the divergene reached about 1.83% between E. cephalophus and the three species of Muntiacus. Using the software of MEGA to analyse molecular phylogeny, Phylogenetic trees were constructed with neighbor-joining method and maximum parsimony method. The result showed Muntiacus, crinifrons is most closely related to muntjak, with reevesi as their sister species. E. cephalophus is in the other genus.
在本研究中，利用赤麂、黑麂、毛冠鹿和斑羚的基因组DNA扩增钾离子通道基因的部分片段。采用直接T-A克隆法将PCR产物连接到pMD18-T载体质粒上。通过菌落PCR鉴定阳性克隆。使用M13-47/RV-M通用引物测定重组克隆的序列，并通过CLUSTALW软件进行比对。三种麂属物种外显子的核苷酸差异为0.90%-1.44%，斑羚与各麂属鹿之间的差异为0.90%-1.26%。这些麂属鹿的内含子核苷酸差异为0%-1.22%，斑羚与三种麂属物种之间的差异约为1.83%。使用MEGA软件分析分子系统发育，采用邻接法和最大简约法构建系统发育树。结果表明，毛冠鹿与赤麂关系最为密切，黑麂是它们的姐妹物种。斑羚属于另一个属。
