caa a22 4500 378858998 CHVBK 20180305123341.0 cr unu---uuuuu 161128e20031107xx s 000 0 eng 10.1515/BC.2003.170 doi (NATIONALLICENCE)gruyter-10.1515/BC.2003.170 Application of the C4'-Alkylated Deoxyribose Primer System (CAPS) in Allele-Specific Real-Time PCR for Increased Selectivity in Discrimination of Single Nucleotide Sequence Variants [Elektronische Daten] [B. Tews, J. Wilhelm, D. Summerer, M. Strerath, A. Marx, P. Friedhoff, A. Pingoud, M. Hahn] This study describes a quantitative real-time PCR-based approach for discrimination of single nucleotide sequence variants, called CAPS (C4 /alkylated primer system). To increase the discrimination potential of DNA polymerases against competing sequence variants of single nucleotides, 3'-terminally modified primers were designed carrying a methyl residue bound to the C4' of the thymidylate deoxyribose. In a model sequence system positional dependencies of modified thymidylate (at -1, -2, -3) were tested for their influence on discrimination. Highest discrimination factors were obtained with the modification at the ultimate 3'-position. In a comparison between Taq and Pwo DNA polymerases, substantial better results were obtained by Taq DNA polymerase. In contrast to conventional PCR methods for discrimination of sequence variants, achieving a maximum discrimination potential of about 20, CAPS is capable of obtaining sequence-specific amplifications of a desired target among discriminated templates with a dynamic range of 1:100. Therefore, CAPS is a method able to quantitatively discriminate two sequence variants only differing in a single base (e.g., SNP alleles or point mutations). The range of applications of this easy to perform, fast and reliable technique reaches from medical diagnostics, transplantation medicine, molecular and cell biology to human genetics. Targeting of SNPs assures a universal exertion of this method, since these markers are gender-independent, highly abundant and ubiquitous. Copyright © 2003 by Walter de Gruyter GmbH & Co. KG Biochemistry nationallicence Molecular biology nationallicence Cellular biology nationallicence Tews B. aut Wilhelm J. aut Summerer D. aut Strerath M. aut Marx A. aut Friedhoff P. aut Pingoud A. aut Hahn M. aut Biological Chemistry Walter de Gruyter 384/10-11(2003-11-07), 1533-1541 1431-6730 384:10-11<1533 2003 384 bchm https://doi.org/10.1515/BC.2003.170 text/html Onlinezugriff via DOI 1 research article jats NATIONALLICENCE 856 40 https://doi.org/10.1515/BC.2003.170 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Tews B. aut NATIONALLICENCE 700 1- Wilhelm J. aut NATIONALLICENCE 700 1- Summerer D. aut NATIONALLICENCE 700 1- Strerath M. aut NATIONALLICENCE 700 1- Marx A. aut NATIONALLICENCE 700 1- Friedhoff P. aut NATIONALLICENCE 700 1- Pingoud A. aut NATIONALLICENCE 700 1- Hahn M. aut NATIONALLICENCE 773 0- Biological Chemistry Walter de Gruyter 384/10-11(2003-11-07), 1533-1541 1431-6730 384:10-11<1533 2003 384 bchm CC0 http://creativecommons.org/publicdomain/zero/1.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-gruyter