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Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Greaves, Ronda / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter


IMPACT FACTOR 2018: 3.638

CiteScore 2018: 2.44

SCImago Journal Rank (SJR) 2018: 1.191
Source Normalized Impact per Paper (SNIP) 2018: 1.205

Online
ISSN
1437-4331
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Volume 42, Issue 1

Issues

Evaluation of a new efficient procedure for single-nucleotide polymorphism genotyping: tetra-primer amplification refractory mutation system-polymerase chain reaction

Naoko Okayama / Kozue Fujimura / Junji Nakamura / Yutaka Suehiro / Yuichiro Hamanaka / Yuji Hinoda
Published Online: 2005-06-01 | DOI: https://doi.org/10.1515/CCLM.2004.004

Abstract

Tetra-primer amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) is a new efficient method for single-nucleotide polymorphism (SNP) genotyping. To determine the optimal conditions for ARMS-PCR we attempted to genotype ten SNPs. DNA was extracted from the peripheral blood of 168 unrelated healthy Japanese volunteers. Two problems inhibited uniform efficiency of the amplification of three bands. The first problem was the lower amplification efficiency of the shorter and allele-specific products compared with the largest product. This phenomenon was overcome by increasing the relative concentration of the inner primers. The second problem was non-specific amplification of the shorter products. To reduce the amplification of these nonspecific bands, adjusting any one of the following PCR conditions was effective: i) reducing the ratio of the inner primer concentration relative to that of the outer primers; ii) increasing the annealing temperature for the initial 5–10 cycles; iii) hot start PCR. With these procedures all ten of the SNPs were successfully genotyped. Our present data may be useful in the further application of tetra-primer ARMS-PCR to SNP genotyping.

About the article

Published Online: 2005-06-01

Published in Print: 2004-02-16


Citation Information: Clinical Chemistry and Laboratory Medicine (CCLM), Volume 42, Issue 1, Pages 13–16, ISSN (Online) 1437-4331, ISSN (Print) 1434-6621, DOI: https://doi.org/10.1515/CCLM.2004.004.

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