人类OPRM1-EXON1的克隆与探针的制备

目的　克隆、测序人类 OPRM1- EXON1,并用非同位素 生物素标记法对该基因进行标记、制备探针,用于OPRM1- EXON1的表达及功能研究。方法　通过PCR法扩增目的基因片段,并将其连接到pGEM- T载体上,转入感受态细胞中进行重组并克隆,经酶切和基因测序进行鉴定,用非同位素 生物素标记法进行探针的标记与制备。结果　经过PCR扩增的目的基因片段大小(2.2 kb)与理论上片段大小一致,经过测序证实其序列与 NCBI提供的序列相同,用此片段成功的制备了用于研究阿片受体基因的探针。结论　从基因组中成功克隆了人类 OPRM1- EX ON1,并制备了探针,为深入研究吗啡依赖相关基因及基因表达创造了必要的条件。

The cloning of human OPRMI-EXON1 and preparation of its probe

Objective To clone and sequence human OPRMI-EXON1, mark it by way of nonisotope-biotin-label, and prepare its probe to study the expression and function of human OPRMI-EXON1. Methods The target gene fragment was amplified by polymerase chain reaction (PCR), and connected to the pGEM-T vector plasmid, then recombined and cloned in competent cell. After that, it was identified by cutting with restriction endonucleases and gene sequence. Finally, we marked it and prepared its probe by nonisotope-biotin-label technique. Results It was demonstrated that the target gene length (2.2kb) amplified by polymerase chain reaction had the same size with the reckoned size in theory and had the same sequence with that of NCBI database. The probe which was used to study the opioid receptor gene was successfully prepared. Conclusion The human OPRMI-EXON1 can be successfully cloned and the probe successfully prepared from the genome, which creates a favorable basis for further research of the morphine-related genes and the expression of their dependence.