title: Separation principles of cycling temperature capillary electrophoresis.
abstract: High throughput means to detect and quantify low-frequency mutations (<10(-2) ) in the DNA-coding sequences of human tissues and pathological lesions are required to discover the kinds, numbers, and rates of genetic mutations that (i) confer inherited risk for disease or (ii) arise in somatic tissues as events required for clonal diseases such as cancers and atherosclerotic plaque.While throughput of linear DNA sequencing methods has increased dramatically, such methods are limited by high error rates (>10(-3) ) rendering them unsuitable for the detection of low-frequency risk-conferring mutations among the many neutral mutations carried in the general population or formed in tissue growth and development. In contrast, constant denaturing capillary electrophoresis (CDCE), coupled with high-fidelity PCR, achieved a point mutation detection limit of <10(-5) in exon-sized sequences from human tissue or pooled blood samples. However, increasing CDCE throughput proved difficult due to the need for precise temperature control and the time-consuming optimization steps for each DNA sequence probed. Both of these problems have been solved by the method of cycling temperature capillary electrophoresis (CTCE). The data presented here provide a deeper understanding of the separation principles involved in CTCE and address several elements of a previously presented two-state transport model.
authors.affiliation: Department of Surgical Oncology and Tumor biology, Radiumhospitalet, Oslo University Hospital, Montebello, Oslo, Norway.
authors.forename: Per Olaf
authors.initials: PO
authors.lastname: Ekstrøm
authors.forename: David J
authors.initials: DJ
authors.lastname: Warren
authors.forename: William G
authors.initials: WG
authors.lastname: Thilly
mesh_terms: D004247:DNA / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D004252:DNA Mutational Analysis / Q000379:methods; D058645:Denaturing Gradient Gel Electrophoresis; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D013696:Temperature
chemical_list: D004247:DNA