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Construction of eukaryotic expression vector of TSLC1 gene
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 hu...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Termedia Publishing House
2011
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3258782/ https://www.ncbi.nlm.nih.gov/pubmed/22291791 http://dx.doi.org/10.5114/aoms.2011.24124 |
Sumario: | 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. |
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