NDLI: Identification of mutations from phenotype-driven ENU mutagenesis in mouse Chromosome 7

Content Provider	Springer Nature Link
Author	Culiat, Cymbeline T. Klebig, Mitchell L. Liu, Zhaowei Monroe, Heidi Stanford, Beverly Desai, Jayashree Tandan, Samvit Hughes, Lori Kerley, Marilyn K. Carpenter, Donald A. Johnson, Dabney K. Rinchik, Eugene M. Li, Qingbo
Copyright Year	2005
Abstract	We have used the new high-throughput mutation-scanning technique temperature-gradient capillary electrophoresis (TGCE) for the identification of point mutations induced by N-ethyl-N-nitrosourea (ENU) in the mouse genome. TGCE detects the presence of heteroduplex molecules formed between a wild-type gene segment and the corresponding homologous segment containing an induced mutation or a naturally occurring single nucleotide polymorphism (SNP). Partially denatured heteroduplex molecules are resolved from homoduplexes by virtue of their differential mobilities during capillary electrophoresis conducted in a finely controlled temperature gradient. Simultaneous heteroduplex analysis of 96 amplicons ranging from 150 to 600 bp in size is achieved in approximately 45 min without the need for predetermining the melting profile of each fragment. Initially, we exploited known mouse mutations to develop TGCE protocols for analyzing unpurified PCR samples amplified from crude tail-DNA preparations. TGCE was then applied to the rapid identification of three new ENU-induced mutations recovered from regional mutagenesis screens of a segment of mouse Chromosome 7. Enzyme assays and quantitative reverse transcription-PCR (qRT-PCR) methods validated these new mutations. Our data demonstrate that rapid mutation scanning with TGCE, followed by sequence verification only of detected positives, is an efficient approach to the identification of point mutations in the mouse genome.
Starting Page	555
Ending Page	566
Page Count	12
File Format	PDF
ISSN	09388990
Journal	Mammalian Genome
Volume Number	16
Issue Number	8
e-ISSN	14321777
Language	English
Publisher	Springer-Verlag
Publisher Date	2005-09-14
Publisher Place	New York
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Anatomy Cell Biology Zoology
Content Type	Text
Resource Type	Article
Subject	Genetics

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1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
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3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
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