| Autor: |
Malik, Muniba Abid Munir, Haider, Muhammad Saleem, Zhai, Ying, Khan, Muhammad Azmat Ullah, Pappu, Hanu R. |
| Předmět: |
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| Zdroj: |
Journal of Plant Diseases & Protection; Feb2023, Vol. 130 Issue 1, p23-33, 11p |
| Abstrakt: |
Mastreviruses are an emerging group of viruses transmitted by leafhoppers and infect both monocot and dicot plants. Chickpea chlorotic dwarf virus (CpCDV), a mastrevirus, is increasingly becoming important in many parts of the world. Various gRNA constructs targeting conserved genomic regions were used singly or in tandem (three constructs together) in CRISPR/Cas9-based approach for triggering resistance against CpCDV. Following inoculation with the infectious clone and each of the constructs, Nicotiana benthamiana was tested for the relative accumulation of the viral DNA through qPCR. Out of five uniplex and three multiplex, one multiplex construct consisting of three genomic regions (LIR, Rep and RepA) showed a significant decrease in viral titre when 2^-∆∆Ct was compared at 7 days after inoculation (dai), 14 dai and 21 dai. When this multiplex construct was agroinfiltrated into the tomato, the qPCR for the viral titre was 0.266 ± 0.06 at 7 dai and 0.069 ± 0.034 when compared with the CpCDV positive control. The expression of gRNA and Cas9 decreased from 7.08 ± 1.24 to 5.22 ± 0.36 and 111.2 ± 5.19 to 75.2 ± 5.26 from 4 to 7 dai, respectively. Positive control plants infiltrated with an infectious clone showed chlorotic leaves, thickened veins and stunted growth at 7 dai, whereas plants infiltrated with the multiplex construct showed mild leaf yellowing and thickened veins and ultimately recovered from the symptoms at 21 dai stage. CRISPR/Cas9-based multiplex of three sgRNAs significantly reduced the viral titre; however, other triplex constructs of sgRNA and individual sgRNA failed to produce consistent results. Our study shows the potential of Cas9-based genome editing technique to generate resistance to CpCDV. [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
| Externí odkaz: |
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