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Open AccessArticle

Knockout of Pi21 by CRISPR/Cas9 and iTRAQ-Based Proteomic Analysis of Mutants Revealed New Insights into M. oryzae Resistance in Elite Rice Line

1
College of Agriculture, State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, Guangxi University, Nanning 530004, China
2
State Key Laboratory for Conservation and Utilization of Subtropical Agricultural Bioresources, South China Agricultural University, Guangzhou 510642, China
*
Authors to whom correspondence should be addressed.
Genes 2020, 11(7), 735; https://doi.org/10.3390/genes11070735
Received: 4 June 2020 / Revised: 24 June 2020 / Accepted: 30 June 2020 / Published: 2 July 2020
(This article belongs to the Special Issue Genetics and Epigenetics of Biotic Stress Response in Plants)
Rice blast (Magnaporthe oryzae) is a devastating disease affecting rice production globally. The development of cultivars with host resistance has been proved to be the best strategy for disease management. Several rice-resistance genes (R) have been recognized which induce resistance to blast in rice but R gene-mediated mechanisms resulting in defense response still need to be elucidated. Here, mutant lines generated through CRISPR/Cas9 based targeted mutagenesis to investigate the role of Pi21 against blast resistance and 17 mutant plants were obtained in T0 generation with the mutation rate of 66% including 26% bi-allelic, 22% homozygous, 12% heterozygous, and 3% chimeric and 17 T-DNA-free lines in T1 generation. The homozygous mutant lines revealed enhanced resistance to blast without affecting the major agronomic traits. Furthermore, comparative proteome profiling was adopted to study the succeeding proteomic regulations, using iTRAQ-based proteomic analysis. We identified 372 DEPs, among them 149 up and 223 were down-regulated, respectively. GO analysis revealed that the proteins related to response to stimulus, photosynthesis, carbohydrate metabolic process, and small molecule metabolic process were up-regulated. The most of DEPs were involved in metabolic, ribosomal, secondary metabolites biosynthesis, and carbon metabolism pathways. 40S ribosomal protein S15 (P31674), 50S ribosomal protein L4, L5, L6 (Q10NM5, Q9ZST0, Q10L93), 30S ribosomal protein S5, S9 (Q6YU81, Q850W6, Q9XJ28), and succinate dehydrogenase (Q9S827) were hub-proteins. The expression level of genes related to defense mechanism, involved in signaling pathways of jasmonic acid (JA), salicylic acid (SA), and ethylene metabolisms were up-regulated in mutant line after the inoculation of the physiological races of M. oryzae as compared to WT. Our results revealed the fundamental value of genome editing and expand knowledge about fungal infection avoidance in rice. View Full-Text
Keywords: rice; M. oryzae; CRISPR/Cas9; Pi21; homozygous; resistance; proteomics; iTRAQ rice; M. oryzae; CRISPR/Cas9; Pi21; homozygous; resistance; proteomics; iTRAQ
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MDPI and ACS Style

Nawaz, G.; Usman, B.; Peng, H.; Zhao, N.; Yuan, R.; Liu, Y.; Li, R. Knockout of Pi21 by CRISPR/Cas9 and iTRAQ-Based Proteomic Analysis of Mutants Revealed New Insights into M. oryzae Resistance in Elite Rice Line. Genes 2020, 11, 735. https://doi.org/10.3390/genes11070735

AMA Style

Nawaz G, Usman B, Peng H, Zhao N, Yuan R, Liu Y, Li R. Knockout of Pi21 by CRISPR/Cas9 and iTRAQ-Based Proteomic Analysis of Mutants Revealed New Insights into M. oryzae Resistance in Elite Rice Line. Genes. 2020; 11(7):735. https://doi.org/10.3390/genes11070735

Chicago/Turabian Style

Nawaz, Gul; Usman, Babar; Peng, Haowen; Zhao, Neng; Yuan, Ruizhi; Liu, Yaoguang; Li, Rongbai. 2020. "Knockout of Pi21 by CRISPR/Cas9 and iTRAQ-Based Proteomic Analysis of Mutants Revealed New Insights into M. oryzae Resistance in Elite Rice Line" Genes 11, no. 7: 735. https://doi.org/10.3390/genes11070735

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