CRISPR/Cas9 mediated high efficiency knockout of the eye color gene Vermillion in Helicoverpa zea (Boddie)

PloS One
Omaththage P PereraCalvin A Pierce

Abstract

Among various genome editing tools available for functional genomic studies, reagents based on clustered regularly interspersed palindromic repeats (CRISPR) have gained popularity due to ease and versatility. CRISPR reagents consist of ribonucleoprotein (RNP) complexes formed by combining guide RNA (gRNA) that target specific genomics regions and a CRISPR associated nuclease (Cas). The gRNA targeting specific gene sequences may be delivered as a plasmid construct that needs to be transcribed or as a synthetic RNA. The Cas nuclease can be introduced as a plasmid construct, mRNA, or purified protein. The efficiency of target editing is dependent on intrinsic factors specific to each species, the target gene sequence, and the delivery methods of CRISPR gRNA and the Cas nuclease. Although intrinsic factors affecting genome editing may not be altered in most experiments, the delivery method for CRISPR/Cas reagents can be optimized to produce the best results. In this study, the efficiency of genome editing by CRISPR/Cas system in the bollworm, Helicoverpa zea (Boddie), was evaluated using ribonucleoprotein (RNP) complexes assembled by binding synthetic gRNA with purified Cas9 nuclease engineered with nuclear localization signals to ...Continue Reading

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Citations

Mar 7, 2020·Insect Molecular Biology·V KoidouJ Vontas
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Datasets Mentioned

BETA
MG976796
MF598173

Methods Mentioned

BETA
electrophoresis
PCR
gene knockouts

Software Mentioned

MEGA
Vector NTI
BlastStation
- Local
Local
Vector NTI Advance
AlignX

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