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

作者：Zhao, Sheng; Zhang, Cuicui; Wang, Liqun; Luo, Minxuan; Zhang, Peng; Wang, Yue; Malik, Waqar Afzal; Chen, Peng; Qiu, Xianjin; Wang, Chongrong; Lu, Hong; Xiang, Yong; Liu, Yuwen; Ruan, Jue; Qian, Qian^*; Zhi, Haijian^*; Chang, Yuxiao^*

来源：Journal of Integrative Plant Biology, 2023, 65(3): 633-645.

DOI：10.1111/jipb.13395

• background selection
• foreground genotyping
• primer-template mismatched annealing
• marker-assisted breeding
• whole-genome genotyping

Whole-genome genotyping methods are important for breeding. However, it has been challenging to develop a robust method for simultaneous foreground and background genotyping that can easily be adapted to different genes and species. In our study, we accidently discovered that in adapter ligation-mediated PCR, the amplification by primer-template mismatched annealing (PTMA) along the genome could generate thousands of stable PCR products. Based 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 PTMA. Unlike 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 species. FBI-seq therefore provides a prolific, robust, and accurate method for simultaneous foreground and background genotyping to facilitate breeding in the post-genomics era.

• 单位
  南京农业大学; 广东省农业科学院