Construction and Identification of Human Tissue Kallikrein Gene Eukaryotic Expressing Vector  

作　　者：戴勇 彭武建 李体远 杜垬 比孙文学 陈德珩 徐卓家 

机构地区：[1]The Second Affiliated Hospital, Medical College ofJinan University （Shenzhen People＇s Hospital）, Shenzhen 518020, China [2]The First Affiliated Hospital of Wenzhou Medical College, Wenzhou 32500, China

出　　处：《Journal of Huazhong University of Science and Technology(Medical Sciences)》2007年第2期164-166,共3页华中科技大学学报（医学英德文版）

基　　金：This project is supported by a program of the 15th Fiveyear Plan of Scientific Research of Bureau of Science and Technology of Guangdong Province (No. 2001c11801)

摘　　要：To clone and sequence the human tissue kallikrein gene of Chinese, and to construct eukaryotic expression recombinant of KK, total RNA was extracted from human pancreas and human tissue kallikrein gene cDNA was amplified by PCR after reverse-transcription by using Oligo（dT） primer. The original kallikrein cDNA was recovered and filled with Klenow enzyme and inserted into KS plasmid. After restriction endonuclease digestion, KK cDNA was sequenced by ABI 377 analyzer. Then the KK gene was amplified from pBluescript KSKK and inserted into pcDNA3. A sequence comparison showed that the cloned kallikrein gene was only one nucleotide different from that reported in the Genbank. The coding amino acid was Asp in the Genbank gene, while the coding amino acid of Chinese kallikrein gene was Asn. The KK cDNA fragment was inserted into the eukaryotic expression vector pcDNA3. The cloned kallikrein gene and the pcDNA3KK can be used for further study in gene therapy.To clone and sequence the human tissue kallikrein gene of Chinese, and to construct eukaryotic expression recombinant of KK, total RNA was extracted from human pancreas and human tissue kallikrein gene cDNA was amplified by PCR after reverse-transcription by using Oligo（dT） primer. The original kallikrein cDNA was recovered and filled with Klenow enzyme and inserted into KS plasmid. After restriction endonuclease digestion, KK cDNA was sequenced by ABI 377 analyzer. Then the KK gene was amplified from pBluescript KSKK and inserted into pcDNA3. A sequence comparison showed that the cloned kallikrein gene was only one nucleotide different from that reported in the Genbank. The coding amino acid was Asp in the Genbank gene, while the coding amino acid of Chinese kallikrein gene was Asn. The KK cDNA fragment was inserted into the eukaryotic expression vector pcDNA3. The cloned kallikrein gene and the pcDNA3KK can be used for further study in gene therapy.

关 键 词：KALLIKREIN GENE cloning VECTOR sequence analysis 

分 类 号：R346[医药卫生—基础医学]