Noureen Azka, Khan Muhammad Zuhaib, Amin Imran, Zainab Tayyaba, Mansoor Shahid
Agricultural Biotechnology Division, National Institute for Biotechnology and Genetic Engineering (NIBGE), Pakistan Institute of Engineering and Applied Sciences (PIEAS), Faisalabad, Pakistan.
University Institute of Biochemistry and Biotechnology (UIBB), Pir Mehr Ali Shah-Arid Agriculture University, Rawalpindi, Pakistan.
Front Genet. 2022 Jul 13;13:922019. doi: 10.3389/fgene.2022.922019. eCollection 2022.
Potato ( L.) is an important staple food around the world, and potato virus Y (PVY) is a major constraint for potato production. The VPg protein of PVY interacts with the translation initiation factor of the host that works as a susceptibility factor during infection. The interaction between eIF4E and VPg was disrupted by CRISPR/Cas9. The homozygous conserved region of of the potato variety "Kruda" was mutated by CRISPR/Cas9. Tracking of insertion, deletion, and conversion events was performed by Sanger sequencing with ∼15% editing efficiency. Truncated and mutated eIF4E proteins were unable to interact with VPg, and the virus was not able to exploit the host machinery for replication and systemic spreading. Mutated lines showed enhanced resistance to PVY strain. DAS-ELISA and RT-PCR were used for validation of the observed resistance. PVY resistance in tetraploid lines CRISPR/Cas9 provides a route to develop novel resistant potato cultivars.
马铃薯（茄属）是全球重要的主食作物，而马铃薯Y病毒（PVY）是马铃薯生产的主要限制因素。PVY的VPg蛋白与宿主的翻译起始因子相互作用，该因子在感染过程中作为易感因子起作用。eIF4E与VPg之间的相互作用被CRISPR/Cas9破坏。马铃薯品种“Kruda”的eIF4E纯合保守区域被CRISPR/Cas9突变。通过Sanger测序对插入、缺失和转换事件进行追踪，编辑效率约为15%。截短和突变的eIF4E蛋白无法与VPg相互作用，病毒无法利用宿主机制进行复制和系统传播。突变的eIF4E品系对PVY毒株表现出增强的抗性。采用双抗夹心酶联免疫吸附测定法（DAS-ELISA）和逆转录-聚合酶链反应（RT-PCR）验证观察到的抗性。四倍体品系中通过CRISPR/Cas9获得的PVY抗性为培育新型抗性马铃薯品种提供了一条途径。
