Construction and expression of human PDX-1 eukaryotic expression vector
- VernacularTitle:人PDX-1基因真核表达载体的构建及表达
- Author:
han-bei, CHEN
;
yan, DONG
;
qing, SU
- Publication Type:Journal Article
- Keywords:
pancreatic duodenal homeobox 1;
gene clone;
eukaryotic expression vector;
transfection
- From:Journal of Shanghai Jiaotong University(Medical Science)
2006;0(01):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct eukaryotic expression vector of human pancreatic duodenal homeobox 1(PDX-1) gene,and to detect its expression in NIH3T3 cell lines. Methods The whole coding sequence of PDX-1 gene was amplified by polymerase chain reaction(PCR) from human pancreatic-cell tumors cDNA.The fragment was inserted into eukaryotic expression vector pcDNA3.1 plasmid.The recombinant plasmid was verified by double digestion and DNA sequencing.The expression of PDX-1 gene in NIH3T3 cells was assayed by Western blot. Results The length of specific fragment amplified by PCR was 852 bp,and the recombinant plasmid pcDNA3.1-PDX-1 showed two bands of 5.5 kb and 852 bp by digestion using respective restriction enzymes BamHⅠand EcoRⅠ.The sequence of PDX-1 gene was approved or confirmed by blasting to GenBank.It was suggested that PDX-1 gene had been cloned into pcDNA3.1 vector correctly.Western blot showed that PDX-1 gene was expressed,which was detected 24 h after pcDNA3.1-PDX-1 plasmid was transfected into NIH3T3 cells. Conclusion The recombinant eukaryotic expression vector pcDNA3.1-PDX-1 was successfully constructed and expressed in NIH3T3 cell lines.
