title: High throughput DNA sequence variant detection by conformation sensitive capillary electrophoresis and automated peak comparison.
abstract: We report the development of a heteroduplex-based mutation detection method using multicapillary automated sequencers, known as conformation-sensitive capillary electrophoresis (CSCE). Our optimized CSCE protocol detected 93 of 95 known base substitution sequence variants. Since the optimization of the method, we have analyzed 215 Mb of DNA and identified 3397 unique variants. An analysis of this data set indicates that the sensitivity of CSCE is above 95% in the central 56% of the average PCR product. To fully exploit the mutation detection capacity of this method, we have developed software, canplot, which automatically compares normal and test results to prioritize samples that are most likely to contain variants. Using multiple fluorescent dyes, CSCE has the capacity to screen over 2.2 Mb on one ABI3730 each day. Therefore this technique is suitable for projects where a rapid and sensitive DNA mutation detection system is required.
authors.affiliation: The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, UK.
authors.forename: Helen
authors.initials: H
authors.lastname: Davies
authors.forename: Ed
authors.initials: E
authors.lastname: Dicks
authors.forename: Philip
authors.initials: P
authors.lastname: Stephens
authors.forename: Charles
authors.initials: C
authors.lastname: Cox
authors.forename: Jon
authors.initials: J
authors.lastname: Teague
authors.forename: Chris
authors.initials: C
authors.lastname: Greenman
authors.forename: Graham
authors.initials: G
authors.lastname: Bignell
authors.forename: Sarah
authors.initials: S
authors.lastname: O'meara
authors.forename: Sarah
authors.initials: S
authors.lastname: Edkins
authors.forename: Adrian
authors.initials: A
authors.lastname: Parker
authors.forename: Claire
authors.initials: C
authors.lastname: Stevens
authors.forename: Andrew
authors.initials: A
authors.lastname: Menzies
authors.forename: Matt
authors.initials: M
authors.lastname: Blow
authors.forename: Bill
authors.initials: B
authors.lastname: Bottomley
authors.forename: Mark
authors.initials: M
authors.lastname: Dronsfield
authors.forename: P Andrew
authors.initials: PA
authors.lastname: Futreal
authors.forename: Michael R
authors.initials: MR
authors.lastname: Stratton
authors.forename: Richard
authors.initials: R
authors.lastname: Wooster
mesh_terms: D000465:Algorithms; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D015894:Genome, Human; D006801:Humans; D009690:Nucleic Acid Conformation; D017354:Point Mutation / Q000235:genetics; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D015203:Reproducibility of Results
chemical_list: D004247:DNA