The cloning and construction of eukaryotic expression vector of human CIDE3 and its proapoptotic effect on HeLa cells
- VernacularTitle:人CIDE3基因克隆和真核表达载体构建及促凋亡作用的研究
- Author:
Shufang WANG
;
Qing LI
;
Li YAO
;
Hong CHENG
;
Jing ZHANG
;
Jing YE
;
Fanfan LI
;
Nanlin LI
- Publication Type:Journal Article
- Keywords:
Cell death-inducing DFF45-like effectors 3;
Gene clone;
Eukaryotic expression vector;
Apoptosis
- From:
Journal of Medical Postgraduates
2003;0(07):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To clone the human cell death-inducing DFF45-like effectors 3(CIDE3) full length cDNA for construction the eukaryotic expression vector pcDNA3.l(+)-CIDE3 and to study its bioactivity.Methods:① Total RNA was extracted from human white adipose tissues and the desired cDNA fragment was obtained by RT-PCR.After the fragment had being inserted into an eukaryotic expression vector pcDNA3.l(+),the resulted recombinant plasmid pcDNA3.l(+)-CIDE3 was transformed into DH5?.The positive clone was selected and confirmed to contain full length of CIDE3 cDNA by agarose gel and DNA sequence analysis.②After the pcDNA3.l(+)-CIDE3 plasmid was transfected into HeLa cells under lipofectamine mediation,the effect of target gene expression on growth of HeLa cells was analysed by TUNEL staining. Results:① The recombinant eukaryotic expression plasmid pcDNA3.l(+)-CIDE3 was constructed successfully and the sequence of CIDE3 was consistent with that of genebank.②After transfected pcDNA3.l(+)-CIDE3,HeLa cells presented distinguished apoptosis(about 15%),compared with that of transfected plasmid pcDNA3.l(+)(
