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[Construction of eukaryotic expression plasmids containing green fluorescent protein gene and CYP19 WT or its variants].
- Source :
-
Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences [Zhejiang Da Xue Xue Bao Yi Xue Ban] 2011 Mar; Vol. 40 (2), pp. 189-94. - Publication Year :
- 2011
-
Abstract
- Objective: To construct eukaryotic expression plasmids containing green fluorescent protein gene and CYP19 wild-type or its variants (W39R, R264C, W39R-R264C) and to observe its expression in MCF-7 and Bcap-37 cells.<br />Methods: The aromatase WT cDNA sequence was obtained by RT-PCR amplification and cloned into the eukaryotic expression vector pcDNA3.1(+). pcDNA3.1(+)-CYP19-GFP plasmid was then used as the template for site-directed mutation to create variant constructs (W39R, R264C, W39R-R264C). pcDNA3.1(+)-CYP19-GFP was transfected and expressed in MCF-7 and Bcap-37 cells.<br />Result: The construction of pcDNA3.1(+)-CYP19-GFP plasmid was confirmed by enzyme digestion and DNA sequencing. pcDNA3.1(+)-CYP19(W39R)-GFP, pcDNA3.1(+)-CYP19(R264C)-GFP, pcDNA3.1(+)- CYP19(W39R-R264C)-GFP plasmids were confirmed by DNA sequencing. The MCF-7 and Bcap-37 cells transfected with the pcDNA3.1(+)-CYP19-GFP plasmid expressed reporter gene of GFP.<br />Conclusion: The eukaryotic expression plasmids have been constructed and expressed in MCF-7 and Bcap-37 cells successfully, which lays the foundation for the research of biological activities of CYP19 variant allozymes.
Details
- Language :
- Chinese
- ISSN :
- 1008-9292
- Volume :
- 40
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences
- Publication Type :
- Academic Journal
- Accession number :
- 21488216
- Full Text :
- https://doi.org/10.3785/j.issn.1008-9292.2011.02.012