【摘要】 目的　设计特异引物克隆大鼠肌细胞生成素 (myogenin)的cDNA。方法　用逆转录聚合酶链反应 (reversetranscriptionpolymerasechainreaction ,RT PCR)合成myogenincDNA ;经提纯后 ,连接到克隆载体pGEM T ,形成重组的克隆载体pGEMT myogenin。pGEMT myogenin转化大肠杆菌Ecoli DH5α内扩增 ;提取、纯化的pGEMT myogenin ,经酶切、PCR鉴定及DNA测序 ,来判断所克隆myogenincDNA的正确性。结果　RT PCR产物经凝胶电泳鉴定 ,含有大小正确的DNA片段 ;提取、纯化的pGEMT myogenin质粒经限制性内切酶酶切、PCR鉴定 ,含有大小正确的DNA片段 ,且该片段含有所设计的酶切位点。DNA测序结果证实所克隆的myogenincDNA序列正确。结论　该研究成功地克隆了myogenincDNA。更多还原

【Abstract】 Objective To clone rat myogenin cDNA by design of specific primers.Methods The myogenin cDNA was composed by reverse transcription polymerase chain reaction (RT PCR).The purified products were ligated with the cloning vecter pGEM T(Promega, USA). The recombinant cloning vector pGEMT myogenin was formed. Then pGEMT moygenin was transformed into Escherichia Coli DH5α for amplifying target gene. After plasmids pGEMT myogenin were extracted and purified, the coding sequence of myogenin cDNA was verified by digestion with restriction endonuclease, PCR checkup and DNA sequencing. Results The RT PCR products was checked by agarose gel electrophoresis analysis. The result confirmed the right length of inserted DNA fragment. The extracted and purified plasmids pGEMT myogenin were digested by restriction endonuclease and checked up by PCR. The result confirmed the right length of inserted DNA fragment with designed digested site. The DNA sequencing result confirmed the cloned myogenin cDNA sequence was right. Conclusion The myogenin cDNA is cloned successfully.更多还原