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A prolific and robust whole-genome genotyping method using PCR amplification via primer-template mismatched annealing

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Journal of Integrative Plant Biology 2023年第3期65卷 633-645页

作者： Sheng Zhao Cuicui Zhang Liqun Wang Minxuan Luo Peng Zhang Yue Wang Waqar Afzal Malik Yue Wang Peng Chen Xianjin Qiu Chongrong Wang Hong Lu Yong Xiang Yuwen Liu Jue Ruan Qian Qian Haijian Zhi Yuxiao Chang National Key Laboratory for Crop Genetics and Germplasm Enhancement National Center for Soybean ImprovementNanjing Agricultural UniversityNanjing 210095China Guangdong Laboratory of Lingnan Modern Agriculture Genome Analysis Laboratory of the Ministry of Agriculture and Rural AffairsAgricultural Genomics Institute at ShenzhenChinese Academy of Agricultural SciencesShenzhen 518120China College of Agriculture Yangtze UniversityJingzhou 434023China Guangdong Provincial Key Laboratory of New Technology in Rice Breeding Rice Research InstituteGuangdong Academy of Agricultural SciencesGuangzhou 510640China

whole-genome genotyping methods are important for ***,it has been challenging to develop a robust method for simultaneous foreground and background genotyping that can easily be adapted to different genes and *** our study,we accidently discovered that in adapter ligation-mediated PCR,the amplification by primertemplate mismatched annealing(PTMA)along the genome could generate thousands of stable PCR *** on this observation,we consequently developed a novel method for simultaneous foreground and background integrated genotyping by sequencing(FBI-seq)using one specific primer,in which foreground genotyping is performed by primer-template perfect annealing(PTPA),while background genotyping employs *** DNA arrays,multiple PCR,or genome target enrichments,FBI-seq requires little preliminary work for primer design and synthesis,and it is easily adaptable to different foreground genes and ***-seq therefore provides a prolific,robust,and accurate method for simultaneous foreground and background genotyping to facilitate breeding in the postgenomics era.

关键词： background selection foreground genotyping primer-template mismatched annealing marker-assisted breeding whole-genome genotyping

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Streamlined whole-genome genotyping through NGS-enhanced thermal asymmetric interlaced(TAIL)-PCR

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Plant Communications 2024年第9期5卷 25-37页

作者： Sheng Zhao Yue Wang Zhenghang Zhu Peng Chen Wuge Liu Chongrong Wang Hong Lu Yong Xiang Yuwen Liu Qian Qian Yuxiao Chang Shenzhen Branch Guangdong Laboratory of Lingnan Modern AgricultureGenome Analysis Laboratory of the Ministry of Agriculture and Rural AffairsAgricultural Genomics Institute at ShenzhenChinese Academy of Agricultural SciencesShenzhen 518120China 2 Guangdong Provincial Key Laboratory of New Technology in Rice Breeding Rice Research InstituteGuangdong Academy of Agricultural SciencesGuangzhou 510640China

whole-genome genotyping(WGG)stands as a pivotal element in genomic-assisted plant ***,sequencing-based approaches for WGG continue to be costly,primarily owing to the high expenses associated with library preparation and the laborious *** prior development of foreground and background integrated genotyping by sequencing(FBI-seq),we discovered that any sequence-specific primer(SP)inherently possesses the capability to amplify a massive array of stable and reproducible non-specific PCR products across the ***,we further improved FBI-seq by replacing the adapter ligated by Tn5 transposase with an arbitrary degenerate(AD)*** protocol for the enhanced FBI-seq unexpectedly mirrors a simplified thermal asymmetric interlaced(TAIL)-PCR,a technique that is widely used for isolation of flanking ***,the improved TAIL-PCR maximizes the primer-template mismatched annealing capabilities of both SP and AD *** addition,leveraging of next-generation sequencing enhances the ability of this technique to assay tens of thousands of genome-wide loci for any *** cost-effective,user-friendly,and powerful WGG tool,which we have named TAIL-PCR by sequencing(TAIL-peq),holds great potential for widespread application in breeding programs,thereby facilitating genome-assisted crop improvement.

关键词： whole-genome genotyping primer-template mismatched annealing specific primer arbitrary degenerate primer molecular breeding

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