Detection of heterozygous nonsense mutations in genes of interest using an Escherichia coli-based stop codon assay
DC Field | Value | Language |
---|---|---|
dc.contributor.author | Moon, Young Joon | - |
dc.contributor.author | Kang, Yoonsung | - |
dc.contributor.author | Choi, Jee-Hye | - |
dc.contributor.author | Lee, Kwang-Ho | - |
dc.date.available | 2019-05-30T06:36:10Z | - |
dc.date.issued | 2007-02 | - |
dc.identifier.issn | 0885-4513 | - |
dc.identifier.issn | 1470-8744 | - |
dc.identifier.uri | https://scholarworks.bwise.kr/cau/handle/2019.sw.cau/24168 | - |
dc.description.abstract | Since nonsense mutations are closely associated with severe conditions of genetic disorders, including familial cancers, rapid and precise detection of those mutations is very important for research purposes and molecular diagnosis. Currently, screening methods such as the FASAY (functional analysis of separated alleles in yeast) and the Y-SC (stop codon assay in yeast) are used for functional detection of nonsense mutations in genes of interest. But these yeast-based approaches are time-consuming, expensive and complicated. In order to circumvent these problems, we, in the present study, devised a novel Escherichia coli-based screening method, the E-SC (E. coli stop codon assay) for the detection of heterozygous nonsense mutations in genes of interest. Our strategy was based on the fact that the plasmid replicating with a low copy number in E. coli allows an effective separation of normal and mutant alleles. Moreover, it relies on the expression vector, resulting in the formation of white and blue colonies for mutant and normal alleles through the expression of PCR-amplified fragment/lacZ fusion protein respectively. To evaluate the applicability of the E-SC method for the detection of the heterozygous truncating mutation, PCR-amplified exon 7 of the StAR [steroidogenic acute regulatory protein; causative gene of the CAH (congenital lipoid adrenal hyperplasia)] and RT (reverse transcription) PCR-amplified fulllength cDNA of MeCP2 (methyl-CpG-binding protein 2; causative gene of Rett syndrome),were used. The ESC showed an almost 1:1 ratio of blue/white colonies in all patients examined, whereas the control samples produced blue colonies only. These results demonstrate that the E-SC system is useful for rapid and precise detection of known and unknown heterozygous truncation mutations in genes that cause genetic disorders and familial cancers. | - |
dc.format.extent | 7 | - |
dc.language | 영어 | - |
dc.language.iso | ENG | - |
dc.publisher | PORTLAND PRESS LTD | - |
dc.title | Detection of heterozygous nonsense mutations in genes of interest using an Escherichia coli-based stop codon assay | - |
dc.type | Article | - |
dc.identifier.doi | 10.1042/BA20060067 | - |
dc.identifier.bibliographicCitation | BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY, v.46, no.2, pp 137 - 143 | - |
dc.description.isOpenAccess | N | - |
dc.identifier.wosid | 000244194600007 | - |
dc.identifier.scopusid | 2-s2.0-33847078425 | - |
dc.citation.endPage | 143 | - |
dc.citation.number | 2 | - |
dc.citation.startPage | 137 | - |
dc.citation.title | BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY | - |
dc.citation.volume | 46 | - |
dc.type.docType | Article | - |
dc.publisher.location | 미국 | - |
dc.subject.keywordAuthor | allele separation | - |
dc.subject.keywordAuthor | stop codon assay | - |
dc.subject.keywordAuthor | Escherichia coli | - |
dc.subject.keywordAuthor | functional analysis of separated alleles in yeast (FASAY) | - |
dc.subject.keywordAuthor | heterozygous nonsense mutation | - |
dc.subject.keywordAuthor | mutation screening | - |
dc.subject.keywordPlus | ACUTE REGULATORY PROTEIN | - |
dc.subject.keywordPlus | TRUNCATING MUTATIONS | - |
dc.subject.keywordPlus | FUNCTIONAL ASSAY | - |
dc.subject.keywordPlus | BREAST-CANCER | - |
dc.subject.keywordPlus | RETT-SYNDROME | - |
dc.subject.keywordPlus | YEAST | - |
dc.subject.keywordPlus | DISEASE | - |
dc.subject.keywordPlus | TUMORS | - |
dc.subject.keywordPlus | BRCA1 | - |
dc.subject.keywordPlus | MECP2 | - |
dc.relation.journalResearchArea | Biochemistry & Molecular Biology | - |
dc.relation.journalResearchArea | Biotechnology & Applied Microbiology | - |
dc.relation.journalWebOfScienceCategory | Biochemistry & Molecular Biology | - |
dc.relation.journalWebOfScienceCategory | Biotechnology & Applied Microbiology | - |
dc.description.journalRegisteredClass | scie | - |
dc.description.journalRegisteredClass | scopus | - |
Items in ScholarWorks are protected by copyright, with all rights reserved, unless otherwise indicated.
84, Heukseok-ro, Dongjak-gu, Seoul, Republic of Korea (06974)02-820-6194
COPYRIGHT 2019 Chung-Ang University All Rights Reserved.
Certain data included herein are derived from the © Web of Science of Clarivate Analytics. All rights reserved.
You may not copy or re-distribute this material in whole or in part without the prior written consent of Clarivate Analytics.