MutExomeSeq Accelerates the Cloning of PmNCA6 Conferring Powdery Mildew Resistance from Triticum boeoticum

Wentao Wan; Renhui Zhao; Peize Zhao; Qiulian Tang; Guofeng Lv; Tiantian Chen; Ling Wang; Shujiang Zang; Ronglin Wu; Zunjie Wang; Shulin Chen; Zongkuan Wang; Xu Zhang; Jinghuang Hu; Hongya Wu; Datong Liu; Yong Zhang; Derong Gao; Hongjie Li; Huagang He; Tongde Bie

doi:10.1016/j.eng.2026.02.027

Engineering ›› :202602027 DOI: 10.1016/j.eng.2026.02.027

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MutExomeSeq Accelerates the Cloning of PmNCA6 Conferring Powdery Mildew Resistance from Triticum boeoticum

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Abstract

Powdery mildew poses a major threat to global wheat production, highlighting the urgent need to identify resistance genes. In this study, we report the cloning of PmNCA6, a powdery mildew resistance gene originating from Triticum boeoticum. Using bulked segregant exome capture sequencing (BSE-Seq) and genetic mapping, we mapped PmNCA6 to a 17-Mb recombination-suppressed interval (680.1-697.1 Mb) on chromosome 7AL. By applying a mutant exome sequencing (MutExomeSeq) approach, we analyzed six ethyl methanesulfonate (EMS)-induced susceptible mutants and identified non-synonymous mutations in a nucleotide-binding leucine-rich repeat (NLR) gene, NLR1. Functional validation through barley stripe mosaic virus-induced gene silencing (BSMV-VIGS) and transgenic complementation confirmed that two alternatively spliced NLR1 transcripts (NLR1_V1 and NLR1_V2) confer resistance to powdery mildew. Phylogenetic analysis revealed that PmNCA6 is orthologous to the stem rust resistance gene Sr22a. Domain-swapping experiments between PmNCA6 and Sr22a demonstrated that the leucine-rich repeat (LRR) domain of PmNCA6 is critical for powdery mildew specificity. Field trials of near-isogenic and recombinant inbred lines (RILs) indicated that PmNCA6-mediated resistance does not compromise yield performance. Screening of 553 Chinese wheat cultivars confirmed the absence of PmNCA6, emphasizing its potential for diversifying resistance sources in breeding programs. This study establishes MutExomeSeq as a robust tool for cloning genes in recombination-suppressed intervals and highlights the potential of engineering synthetic NLRs with tailored LRR domains to combat evolving pathogens.

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PmNCA6 / Powdery mildew resistance / Triticum boeoticum / Wheat breeding / MutExomeSeq

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Wentao Wan, Renhui Zhao, Peize Zhao, Qiulian Tang, Guofeng Lv, Tiantian Chen, Ling Wang, Shujiang Zang, Ronglin Wu, Zunjie Wang, Shulin Chen, Zongkuan Wang, Xu Zhang, Jinghuang Hu, Hongya Wu, Datong Liu, Yong Zhang, Derong Gao, Hongjie Li, Huagang He, Tongde Bie. MutExomeSeq Accelerates the Cloning of PmNCA6 Conferring Powdery Mildew Resistance from Triticum boeoticum. Engineering 202602027 DOI:10.1016/j.eng.2026.02.027

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