Scanning the beta-globin gene for mutations in large populations by denaturing capillary and gel electrophoresis

	Scanning the beta-globin gene for mutations in large populations by denaturing capillary and gel electrophoresis
	Li-Sucholeiki, XC; Hu, GX; Perls, T; Tomita-Mitchell, A; Thilly, WG
刊名	ELECTROPHORESIS
	2005
卷号	26 期号:13 页码:2531-2538
关键词	denaturant capillary electrophoresis denaturing gradient gel electrophoresis beta-globin gene mutation scan
通讯作者	Thilly, WG (reprint author), MIT, Biol Engn Div, 77 Massachusetts Ave,Room 16-743, Cambridge, MA 02139 USA.,thilly@mit.edu
英文摘要	Separation of mutant from nonmutant DNA sequences of similar to 100bp may be accomplished by using defined denaturing conditions of chemical denaturant and/or elevated temperature during electrophoresis on either polyacrylamide slab gels (denaturing gradient gel electrophoresis, DGGE) or capillary gels (constant denaturant capillary electrophoresis, CDCE. In analysis of mutant directly from a polymerase chain reaction (PCR) product mixture, both have detection sensitivities of approximately 1%. CDCE that facilitates an intermediate mutant enrichment step permits detection of mutants at fractions as low as 2 x 10(-6). Here we report the successful application of both approaches to scan for mutations of the human P-globin gene (HBB) in two human population samples of approximately 5000 persons in the HBB. Using DGGE, the coding region and flanking intronic splice sites of HBB were scanned in a population of 4949 Han Chinese individuals in pool sizes of 48 individual DNA samples. Four point mutations ranging in mutant frequency from 0.5 to 0.0002 were identified. Using CDCE with a mutant enrichment step, these same sequences were scanned in a population of 5028, predominantly African-American juveniles (< 9 years) as a single pooled DNA sample. Three point mutations were identified ranging in mutant frequency from 0.13 to 0.0005. This study shows that both the DGGE/small pool and the CDCE/large pool approaches offer the means to define the fine structure map of genetic variation in large population samples, and with appropriately engineered facilities to provide high throughput, should be useful in pangenomic scans to discover genes carrying casual mutations for common diseases.
学科主题	Biochemistry & Molecular Biology; Chemistry
类目[WOS]	Biochemical Research Methods ; Chemistry, Analytical
关键词[WOS]	HUMAN MITOCHONDRIAL-DNA ; THALASSEMIA MUTATIONS ; POINT MUTATIONS ; GENOMIC DNA ; POLYMORPHISMS ; SPECTROMETRY ; DISEASE ; SUBSTITUTIONS ; VARIANTS ; ALLELES
收录类别	SCI
语种	英语
WOS记录号	WOS:000230529600006
内容类型	期刊论文
版本	出版稿
源URL	[http://202.127.25.143/handle/331003/1971]
专题	上海生化细胞研究所_上海生科院生化细胞研究所
推荐引用方式 GB/T 7714	Li-Sucholeiki, XC,Hu, GX,Perls, T,et al. Scanning the beta-globin gene for mutations in large populations by denaturing capillary and gel electrophoresis[J]. ELECTROPHORESIS,2005,26(13):2531-2538.
APA	Li-Sucholeiki, XC,Hu, GX,Perls, T,Tomita-Mitchell, A,&Thilly, WG.(2005).Scanning the beta-globin gene for mutations in large populations by denaturing capillary and gel electrophoresis.ELECTROPHORESIS,26(13),2531-2538.
MLA	Li-Sucholeiki, XC,et al."Scanning the beta-globin gene for mutations in large populations by denaturing capillary and gel electrophoresis".ELECTROPHORESIS 26.13(2005):2531-2538.