Abstract
Begomoviruses infect many plant species including economically important crops worldwide.Conventional approaches for resistance management have not been successful to tackle the emerging and rapidly evolving plant viruses.The last decade has witnessed an unprecedented progress in plant genome editing technologies.CRISPR-Cas9 based gene editing has been successfully applied to engineer plants with tolerance against plant viruses.In the present study, we successfully demonstrated the use of the CRISPR-Cas9 system to suppress disease symptoms of cotton leaf curl disease (CLCuD) caused by begomoviruses.A unique gRNA specific for a consensus DNA sequence of the three most prevalent strains of Cotton Leaf Curl Virus (CLCuV) was synthesized.The consensus sequence was located in the Common Region of CLCuV genome.Transient assays of CLCuV infectious clones (CLCuKoV/CLCuMuB) and Cas9-gRNA construct were performed in 3-4 weeks old Nicotiana benthamiana plants by agroinfiltration.Virus accumulation in local and systemic leaves was analyzed by semi-qPCR which demonstrated lowering of virus titer by 65-75%.Infiltrated plants showed reduced symptom severity relative to control plants infiltrated with virus only.Double strand break (DSB) in DNA at the target sequence mediated by Cas9-gRNA were confirmed by T7E1 assays showing indel mutations in the CLCuV genome vs. no mutations in control plants.The results demonstrated potential use of the CRISPR-Cas9 system for engineering virus resistance in plants.Furthermore, our results also revealed that by utilizing a consensus sequence of multiple viruses, resistance to multiple virus infections can be engineered by a single gene construct.
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Claims 5
Claim · model-written summary
The Cas9-gRNA construct lowered virus titer by 65-75% compared with control plants infiltrated with virus only.
- Taxon
- Nicotiana benthamiana
Nicotiana benthamianae0 - Treatment
- Cas9-gRNA constructe0
- Interactor
- Cotton Leaf Curl VirusCotton leaf curl viruse4pathogen
- Outcome
- virus titere1
- Direction
- loweringe1
- Value
- 65-75%e1
- Comparator
- control plants infiltrated with virus onlye2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transient assays of CLCuV infectious clones (CLCuKoV/CLCuMuB) and Cas9-gRNA construct were performed in 3-4 weeks old Nicotiana benthamiana plants by agroinfiltration”
e1“Virus accumulation in local and systemic leaves was analyzed by semi-qPCR which demonstrated lowering of virus titer by 65-75%”
e2“Infiltrated plants showed reduced symptom severity relative to control plants infiltrated with virus only”
e4“A unique gRNA specific for a consensus DNA sequence of the three most prevalent strains of Cotton Leaf Curl Virus (CLCuV) was synthesized”
Claim · model-written summary
Infiltration with the Cas9-gRNA construct reduced symptom severity relative to control plants infiltrated with virus only.
- Taxon
- Nicotiana benthamianaNicotiana benthamianae0
- Treatment
- Cas9-gRNA constructe0
- Interactor
- Cotton Leaf Curl VirusCotton leaf curl viruse4pathogen
- Outcome
- symptom severitye2
- Direction
- reducede2
- Comparator
- control plants infiltrated with virus onlye2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transient assays of CLCuV infectious clones (CLCuKoV/CLCuMuB) and Cas9-gRNA construct were performed in 3-4 weeks old Nicotiana benthamiana plants by agroinfiltration”
e2“Infiltrated plants showed reduced symptom severity relative to control plants infiltrated with virus only”
e4“A unique gRNA specific for a consensus DNA sequence of the three most prevalent strains of Cotton Leaf Curl Virus (CLCuV) was synthesized”
Claim · model-written summary
Cas9-gRNA induced indel mutations in the CLCuV genome, with no mutations in control plants.
- Taxon
- Nicotiana benthamianaNicotiana benthamianae0
- Interactor
- CLCuVCotton leaf curl viruse3pathogen
- Outcome
- indel mutationse3
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Transient assays of CLCuV infectious clones (CLCuKoV/CLCuMuB) and Cas9-gRNA construct were performed in 3-4 weeks old Nicotiana benthamiana plants by agroinfiltration”
e3“showing indel mutations in the CLCuV genome vs. no mutations in control plants”
Claim · model-written summary
CRISPR-Cas9 has potential use for engineering virus resistance in plants (author interpretation).
- Taxon
- plantsViridiplantaee5
- Outcome
- engineering virus resistancee5
Verbatim
e5“The results demonstrated potential use of the CRISPR-Cas9 system for engineering virus resistance in plants”
Claim · model-written summary
A consensus sequence of multiple viruses can engineer resistance to multiple virus infections with a single gene construct (author interpretation).
- Taxon
- plantsViridiplantaee5
- Outcome
- resistance to multiple virus infectionse6
- Qualifier
- by a single gene constructe6
Verbatim
e5“The results demonstrated potential use of the CRISPR-Cas9 system for engineering virus resistance in plants”
e6“by utilizing a consensus sequence of multiple viruses, resistance to multiple virus infections can be engineered by a single gene construct”
The same questions in other papers
loss of function · Cotton leaf curl virus · Nicotiana benthamianaResults agree
1 paper in the same direction as this paper
- positive
Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.