High-throughput primer design by scoring in piecewise logistic model for multiple polymerase chain reaction variants

Author:

Zeng Huaping,Chen Kexin,Ma Chouxian,Zhu Biyin,Chuan Jun,Zhang Shuan,Tang Lin,Yang Ting,Sun Zhaohui,Yang Xingkun,Wang Yu

Abstract

AbstractPolymerase chain reaction (PCR) variants requiring specific primer types are widely used in various PCR experiments, including generic PCR, inverse PCR, anchored PCR, and ARMS PCR. Few tools can be adapted for multiple PCR variants, and many tools select primers by filtration based on the given parameters, which result in frequent design failures. Here we introduce PrimerScore2, a robust high-throughput primer design tool that can design primers in one click for multiple PCR variants. It scores primers using a piecewise logistic model and the highest-scored primers are selected avoiding the issue of design failure and the necessity to loosen parameters to redesign, and it creatively evaluates specificity by predicting the efficiencies of all target/non-target products. To assess the prediction accuracy of the scores and efficiencies, two next generation sequencing (NGS) libraries were constructed—a 12-plex and a 57-plex—and the results showed that 17 out of 19 (89.5%) low-scoring pairs had a poor depth, 18 out of 19 (94.7%) high-scoring pairs had a high depth, and the depth ratios of the products were linearly correlated with the predicted efficiencies with a slope of 1.025 and a coefficient of determination (R2) 0.935. 116-plex and 114-plex anchored PCR panels designed by PrimerScore2 were applied to 26 maternal plasma samples with male fetuses, the results showed that the predicted fetal DNA fractions were concordant with fractions measured in gold standard method (Y fractions). PrimerScore2 was also used to design 77 monoplex Sanger sequencing primers, the sequencing results indicated that all the primers were effective.

Publisher

Springer Science and Business Media LLC

Subject

Multidisciplinary

Cited by 1 articles. 订阅此论文施引文献 订阅此论文施引文献,注册后可以免费订阅5篇论文的施引文献,订阅后可以查看论文全部施引文献

同舟云学术

1.学者识别学者识别

2.学术分析学术分析

3.人才评估人才评估

"同舟云学术"是以全球学者为主线,采集、加工和组织学术论文而形成的新型学术文献查询和分析系统,可以对全球学者进行文献检索和人才价值评估。用户可以通过关注某些学科领域的顶尖人物而持续追踪该领域的学科进展和研究前沿。经过近期的数据扩容,当前同舟云学术共收录了国内外主流学术期刊6万余种,收集的期刊论文及会议论文总量共计约1.5亿篇,并以每天添加12000余篇中外论文的速度递增。我们也可以为用户提供个性化、定制化的学者数据。欢迎来电咨询!咨询电话:010-8811{复制后删除}0370

www.globalauthorid.com

TOP

Copyright © 2019-2024 北京同舟云网络信息技术有限公司
京公网安备11010802033243号  京ICP备18003416号-3