Problem With Microsatellite Polymerphism Analysis and its Practical Solution Detection of Complex STR Alleles by Single-Strand Conformation Polymorphism (SSCP) Analysis and Denaturing Gradient Gel Electrophoresis (DGGE)

微卫星多态性分析的问题及其通过单链构象多态性 (SSCP) 分析和变性梯度凝胶电泳 (DGGE) 检测复杂 STR 等位基因的实用解决方案

基本信息

  • 批准号:
    09670444
  • 负责人:
  • 金额:
    $ 0.64万
  • 依托单位:
  • 依托单位国家:
    日本
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
  • 财政年份:
    1997
  • 资助国家:
    日本
  • 起止时间:
    1997 至 1998
  • 项目状态:
    已结题

项目摘要

We amplified the complex short tandem repeat (STR) loci D11S488 and D11S554, electrophoresed amplicons on denaturing polyacrylamide gels, and detected them by Southern blotting. Sequencing of 262 bp-long D11S488 alleles revealed five different alleles with structural variations. Sequence analysis of the D115554 locus showed that 237 bp, 225 bp and 217 bp alleles had three, two, and two different sequences, respectively.Single-strand conformation polymorphism (SSCP) analysis of D115488 amplicons on a 6% native polyacrylamide gel at 4゚C enabled us to distinguish four alleles of the same length. Similarly, we were able to identify different D11S554 alleles of the same length by electrophoresis on a 5% polyacrylamide gel at 15゚C.We successfully distinguished four different D115488 alleles (262 bp) by 20-50% denaturing gradient gel electrophoresis (DGGE), and all of the different D115554 alleles mentioned above by 25-35% DGGE.However, some alleles of different lengths showed the same mobility.We applied the SSCP technique to two normal trio cases of disputed parentage, in which four STR loci including D115488 were tested for. Both trios had an apparently identical 262 bp D115488 allele on a denaturing gel. In one case, SSCP analysis increased the combined probability of paternity, ' because the child and the alleged father shared a rare allele. In the other case, the child shared the allele with the mother, but not with the alleged father, which excluded paternity. As a result, paternity was excluded by three of the four loci tested.Allowing simple and rapid detection of sequence polymorphisms, our method is suited for practical use.
对D11 S488和D11 S554两个STR基因座进行扩增,变性聚丙烯酰胺凝胶电泳,Southern印迹检测。262 bp长的D11 S488等位基因的测序显示,5个不同的等位基因的结构变异。D115554基因座的237 bp、225 bp和217 bp等位基因分别有3个、2个和2个不同的序列,D115488扩增产物在6%聚丙烯酰胺凝胶上的单链构象多态性(SSCP)分析表明,D115554基因座有4个相同长度的等位基因。类似地,我们能够通过在15 ℃下在5%聚丙烯酰胺凝胶上电泳来鉴定相同长度的不同D11 S554等位基因。我们通过20-50%变性梯度凝胶电泳(DGGE)成功区分了四种不同的D115488等位基因(262 bp),并且通过25- 35%DGGE区分了上述所有不同的D115554等位基因。本文应用SSCP技术对两例正常的亲子关系有争议的三人组进行了包括D115488在内的4个STR基因座的检测。两个三人组在变性凝胶上具有明显相同的262 bp D115488等位基因。在一个案例中,SSCP分析增加了父子关系的综合概率,因为孩子和所谓的父亲共享一个罕见的等位基因。在另一种情况下,孩子与母亲共享等位基因,但与所谓的父亲不同,这排除了父子关系。结果表明,在所检测的4个位点中,有3个位点排除了亲子关系,该方法简单、快速,适合于实际应用。

项目成果

期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Fukuda,M.: "Subtyping of DllS488 STR alleles by single-strand conformation polymorphism (SSCP) analysis in two cases of disputed parentage" Jpn J Legal Med. 52・1. 42-45 (1998)
Fukuda, M.:“通过单链构象多态性 (SSCP) 分析对两个有争议的亲子关系案例进行 DllS488 STR 等位基因的分型” Jpn J Legal Med 52・1 (1998)。
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    0
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  • 通讯作者:
Fukuda, M.: "Subtyping of D11S488 STR allelesby single-strand conformation polymorphism (SSCP) analysis in two cases of disputed parentage" Jpn J Legal Med. 52 (1). 42-45 (1998)
Fukuda, M.:“在两个有争议的亲子关系案例中通过单链构象多态性 (SSCP) 分析对 D11S488 STR 等位基因进行亚型分析”Jpn J Legal Med。
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    0
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Fukuda,M., Tamaki,Y.: "Subtyping of D11S488 STR alleles by single-strand comformation polymorphism(SSCP)analysis in two cases of disputed parentage" Jpn J Legal Med. 52・1(印刷中). (1998)
Fukuda, M., Tamaki, Y.:“通过单链构象多态性 (SSCP) 分析对两个有争议的亲子关系案例进行 D11S488 STR 等位基因的亚型分析”Jpn J Legal Med 52・1(出版中)。
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  • 影响因子:
    0
  • 作者:
  • 通讯作者:
Fukuda,M.: "Subtyping of D11S488 STR alleles by single-strand conformation polymorphism(SSCP)analysis in two cases of disputed parentage" Jpn J Legal Med. 52・1. 42-45 (1998)
Fukuda, M.:“通过单链构象多态性 (SSCP) 分析对两个有争议的亲子关系案例进行 D11S488 STR 等位基因的亚型分析” Jpn J Legal Med 52・1 (1998)。
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  • 发表时间:
  • 期刊:
  • 影响因子:
    0
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  • 通讯作者:
Fukuda, M.: "Subtyping of complex STR alleles by single-strand conformation polymorphism (SSCP) analysis and denaturing gradient gel electrophoresis (DGGE)" Jpn J Legal Med. 53 (Suppi) (in press).
Fukuda, M.:“通过单链构象多态性 (SSCP) 分析和变性梯度凝胶电泳 (DGGE) 对复杂 STR 等位基因进行亚型分型”Jpn J Legal Med。
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FUKUDA Masako其他文献

FUKUDA Masako的其他文献

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{{ truncateString('FUKUDA Masako', 18)}}的其他基金

Development of a metabolomics-based detection method of filarial infective larvae from blood-sucking insects
基于代谢组学的吸血昆虫丝虫感染性幼虫检测方法的建立
  • 批准号:
    19K06421
  • 财政年份:
    2019
  • 资助金额:
    $ 0.64万
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Discovery of zoonotic filariases derived from wild animals with the use of blood-sucking insects as vectors
以吸血昆虫为载体发现源自野生动物的人畜共患丝虫病
  • 批准号:
    15K07781
  • 财政年份:
    2015
  • 资助金额:
    $ 0.64万
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Research on Occupational Gender Segregation
职业性别隔离研究
  • 批准号:
    14594030
  • 财政年份:
    2002
  • 资助金额:
    $ 0.64万
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)

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