An automated fluorescent single strand conformation polymorphism technique for high throughput mutation screening
10.3760/j.issn:0366-6999.2001.11.007
- VernacularTitle:适用于规模性基因突变扫描的荧光标记单链构象长度多态性检测方法分析
- Author:
Jianping WENG
1
;
Berglund ANNA
;
C.Groop LEIF
;
C.Groop LEIF
Author Information
1. the First Affiliated Hospital Sun Yet San University of Medical Sciences
- Keywords:
genetics;
mutation;
single strand conformation polymorphism;
electrophoresis;
fluorescent label
- From:
Chinese Medical Journal
2001;114(11):1147-1150
- CountryChina
- Language:Chinese
-
Abstract:
Objective To develop a high throughput mutational detection method by mutiple fluorescence-labeled polymerase chain reaction(PCR)products.Methods A total of 27 known mutations including 22 substitutions,3 insertions(1,2 and 7 bp)and 2 deletions(1 and 2 bp)in the hepatocyte nuclear factor(HNF)-4α,glucokinase and HNF-1α genes were tested.During nested PCR,amplified fragments were labeled with three fluorescent dyes.PCR products were visualized with an ABI-377 fluorescence sequencer using 5% glycerol or 10% sucrose in nondenaturing gel conditions.Results Twenty-five of 27 variants(93%)could be detected by combining 5% glycerol and 10% sucrosegel matrix conditions.Twenty-two of 27(82%)and 18 of 27(67%)variants were identified using 5%glycerol and 10% sucrose conditions,respectively.Conclusion This fluorescence-based PCR single strand conformation polymorphism technique represents a simple,non-hazardous,time-saving and sensitive method for high throughput mutation detection.
