Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
Regular Article
Impact of Bridged Nucleic Acid Positions within Blocking Oligonucleotides on DNA Amplification Inhibition in Wild-Type Blocking PCR
Takuma YamashitaYoshinori TsukumoTakenori YamamotoEriko UchidaTokuyuki YoshidaYasunori UchidaTakao Inoue
Author information
JOURNAL OPEN ACCESS FULL-TEXT HTML
Supplementary material

2025 Volume 48 Issue 5 Pages 606-612

Details
Abstract

Detecting low-frequency genetic mutations is crucial for genetic testing, especially in cancer diagnostics. Wild-type blocking PCR identifies these genetic mutations using a blocking oligonucleotide that is fully complementary to wild-type DNA. The blocking oligonucleotide selectively binds to wild-type DNA, inhibiting its amplification by DNA polymerase and allowing preferential amplification of mutant DNA. Bridged nucleic acids (BNAs), with high binding affinities for cDNA, are often incorporated into the blocking oligonucleotide to enhance inhibition. However, the effects of BNA positioning within the blocking oligonucleotide on wild-type DNA amplification inhibition are poorly understood. To address this issue, we evaluated the effects of different BNA positions on amplification inhibition efficacy by comparing blocking oligonucleotides with varying numbers of BNAs at the 5′ end, 3′ end, and central region. Results indicated that BNAs at the 5′ end enhanced the inhibition efficacy, whereas BNAs at the 3′ end notably diminished the inhibition efficacy. Likewise, increasing the number of BNAs in the central region generally decreased the inhibition efficacy. This is one of the first studies to report the importance of BNA positioning in the amplification inhibition efficacy of blocking oligonucleotides.

Fullsize Image
Content from these authors
© 2025 Author(s).
Published by The Pharmaceutical Society of Japan

This article is licensed under a Creative Commons [Attribution-NonCommercial 4.0 International] license.
https://creativecommons.org/licenses/by-nc/4.0/
Previous article Next article
feedback
Top