| 运用重叠延伸PCR技术构建在甘丙肽全长cDNA嵌入第二内含子的重构分子 |
| |
| 引用本文： | 雷小春,司苏晋,薛亮亮,刘田福.运用重叠延伸PCR技术构建在甘丙肽全长cDNA嵌入第二内含子的重构分子[J].生物技术通报,2010(4). |
| |
| 作者姓名： | 雷小春 司苏晋 薛亮亮 刘田福 |
| |
| 作者单位： | 山西医科大学实验动物中心,太原,030001 |
| |
| 基金项目： | 山西省实验动物专项基金 |
| |
| 摘 要： | 构建嵌入第二内含子的甘丙肽(Galanin,GAL)全长基因组cDNA的重构分子。通过RT-PCR扩增出cDNA编区的序列,分别从基因组中扩增出cDNA的5′和3′端部分非编码序列;使用重叠延伸PCR(overlap extention PCR,OE-PCR)方法将三个片段重叠获得全长cDNA序列;再将全长cDNA从第三外显子第15个碱基处分成两部分,分开的cDNA前半部分和后半部分以及第二内含子进行重叠延伸获得重构分子,含有第二内含子的甘丙肽(GAL)全长基因组cDNA;将重构分子连入pMDI9-Tsimple载体。电泳分析观察到清晰的重构分子片段;测序显示重构分子由所设计的序列组成,第二内含子插入的位置准确,且无移码。使用重叠延伸PCR能够成功在cDNA中插入内含子获得一段重构基因。
|
| 关 键 词： | 重叠延伸PCR 甘丙肽 内含子 重构分子 |
Construction of Recombinant Molecule of Galanin Full Length cDNA Including the Second Intron by OE-PCR |
| |
| Lei Xiaochun,Si Sujin,Xue Liangliang,Liu Tianfu.Construction of Recombinant Molecule of Galanin Full Length cDNA Including the Second Intron by OE-PCR[J].Biotechnology Bulletin,2010(4). |
| |
| Authors: | Lei Xiaochun Si Sujin Xue Liangliang Liu Tianfu |
| |
| Institution: | Lei Xiaochun Si Sujin Xue Liangliang Liu Tianfu(Labrotory Animal Centrol of Shanxi Medical University,Taiyuan 030001) |
| |
| Abstract: | The study was design to construct a recombinant gene composed of galanin(GAL)full-length cDNA and the second intron of the galanin gene.Firstly,by RT-PCR the coding cDNA sequence was amplified,and the 5' and 3' noncoding sequences of Gal cDNA was amplified by PCR.Secondly,the full-length cDNA sequence was obtained by overlap extention PCR(OE-PCR).Thirdly,the full-length cDNA was cuted into two part at the 15 bp of the third exon and amplified them respectively.Finally,Using the two part of the full-length c... |
| |
| Keywords: | Overlap extention PCR Galanin Intron Reconstructed molecule |
| 本文献已被 CNKI 万方数据 等数据库收录！ |
|
