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Construction of eukaryotic expression vector of TSLC1 gene.


ABSTRACT: INTRODUCTION:To construct a eukaryotic expression vector of the tumour suppressor in lung cancer 1 (TSLC1) gene, so as to explore the mechanisms of tumour suppression of the gene theoretically. MATERIAL AND METHODS:The open reading frame (ORF) of TSLC1 gene was amplified with RT-PCR from normal human foreskin acrobystia, and cloned to pMD19-T simple vector (TA Clone method). The resultant plasmid was transformed into Escherichia coli JM109 for amplification. The TA Clone recombinant was digested by double restriction enzyme (Bgl II/EcoR I) and analysed with agarose gel electrophoresis. The positive one was sequenced. The inserted DNA fragment was recovered, and then it was mounted into the eukaryotic expression vector pIRES2-EGFP, transformed into E. coli JM109 for amplification. A positive recombinant plasmid named pIRES2-EGFP-TSLC1 was confirmed by Bgl II/EcoR I double-enzyme digestion analysis. RESULTS:RT-PCR amplified the ORF of the TSLC1 gene. It was approximately 1400 base pairs. The obtained DNA was confirmed a high degree of homology with the sequence of TSLC1 cDNA sequence (AY358334) stored at GenBank. CONCLUSIONS:Construction of a TSLC1 eukaryotic expression vector was successful, and it has established a solid foundation for further study.

SUBMITTER: Liang QL 

PROVIDER: S-EPMC3258782 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Construction of eukaryotic expression vector of TSLC1 gene.

Liang Qi-Lian QL   Wang Bi-Rong BR   Li Zhou-Yu ZY   Chen Guo-Qiang GQ   Zhou Yuan Y  

Archives of medical science : AMS 20110801 4


<h4>Introduction</h4>To construct a eukaryotic expression vector of the tumour suppressor in lung cancer 1 (TSLC1) gene, so as to explore the mechanisms of tumour suppression of the gene theoretically.<h4>Material and methods</h4>The open reading frame (ORF) of TSLC1 gene was amplified with RT-PCR from normal human foreskin acrobystia, and cloned to pMD19-T simple vector (TA Clone method). The resultant plasmid was transformed into Escherichia coli JM109 for amplification. The TA Clone recombina  ...[more]

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