The effect of gene polymorphism in promoter and intron 1 on human ApoA I expression.
- Author:
Ai-juan CHENG
1
;
Yong-min MAO
;
Rang-zhuang CUI
Author Information
- Publication Type:Journal Article
- MeSH: Apolipoprotein A-I; genetics; metabolism; Cell Line, Tumor; Cholesterol, HDL; metabolism; Genetic Vectors; Humans; Introns; Luciferases; genetics; metabolism; Point Mutation; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; Recombinant Fusion Proteins; genetics; metabolism; Transfection
- From: Chinese Journal of Medical Genetics 2006;23(6):610-613
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct pGL2-ApoA I luciferase reporter vector containing ApoA I gene regulation area, and to investigate the effect of G --> A and C --> T substitution in ApoA I promoter -75 bp and intron 1 +83 bp region respectively on ApoA I gene expression.
METHODSHuman chromosome DNA fragments containing ApoA I gene were amplified by PCR, and the DNA fragments consisting of ApoA I AA/CC, GG/TT and GG/CC genetypes were selected separately, then pUC vector including above three different DNA fragments was constructed. After digesting pUC vector with Sac I and Bgl II, ligate the different DNA fragments to basic pGL2 vector that containing luciferase reporter gene. Recombinant and PRL-null vector were cotransfected into HepG2 cells by using cationic liposome method. Cells were cultured for 48 h, activity of firefly and renills luciferase was measured.
RESULTSThree vectors with pGL2-ApoA I-L(-2500 to +289 bp) long fragment vectors and 3 with pGL2-ApoA I-S(-145 to +289 bp) short fragment vectors were combinated successfully. Relative activity of luciferase for ApoA I AA/CC or GG/TT was lower than that for GG/CC significantly.
CONCLUSION-75 bp G --> A and +83 bp C --> T substitution in ApoA I gene may inhibit ApoA I gene transcription and expression. It may be the reason why subjects containing -75 bp A and +83 bp T have lower high density lipoprotein cholesterol (HDL-C) concentration.