CRISPR/Cas‐mediated gene editing using purified protein in <i><scp>D</scp>rosophila suzukii</i>

Kalajdzic, Predrag; Schetelig, Marc F.

doi:10.1111/eea.12599

Cited by 32 publications

(31 citation statements)

References 65 publications

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“…In this paper, we describe detailed methods for inducing heritable mutations in the w gene in three tephritid species from three different genera using the CRISPR-Cas9 targeted mutagenesis system. CRISPR mutants have been previously generated in other tephritid species such as B. tryoni and C. capitata (Choo et al, 2018;Meccariello et al, 2017) in addition to other Diptera such as Drosophila melanogaster, D. suzukii, Aedes aegypti and several species of Anopheles mosquitos (Bassett and Liu, 2014;Kistler et al, 2015;Kalajdzic and Schetelig, 2017;Li and Handler, 2017;Dong et al, 2018;Li et al, 2018), but this is the first study that demonstrates these methods in the genus Anastrepha and the species B. dorsalis. The mutations created in these species were caused by nonhomologous end join repair where mutations form spontaneously as a consequence of imperfect DNA repair mechanisms as opposed to homology-directed repair, which requires a template to make specific mutations.…”

Section: Discussionmentioning

confidence: 99%

The ABCs of CRISPR in Tephritidae: developing methods for inducing heritable mutations in the genera Anastrepha, Bactrocera and Ceratitis

Sim

Kauwe

Ruano

et al. 2019

Insect Molecular Biology

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Tephritid fruit flies are destructive agricultural pests that are the targets of expensive population eradication and suppression efforts. Genetic pest management is one of the strategies for reducing or eliminating tephritid populations, relying upon the genetic manipulation of insects to render them sterile or capable of transmitting deleterious traits through gene drive. Currently, radiation, chemical mutagenesis, and transgenic techniques are employed to generate agents for genetic pest management, but new methods must be explored and developed for all tephritid pest species. Targeted mutagenesis induced by nonhomologous end join repair of clustered regularly interspaced short palindromic repeats and the CRISPR associated protein 9 (Cas9) (commonly known as CRISPR/Cas9) has been demonstrated to be an efficient method for creating knock‐out mutants and can be utilized to create germline mutations in Tephritidae. In this paper, we describe detailed methods to knockout the white gene in three tephritid species in the genera Anastrepha, Bactrocera and Ceratitis, including the first demonstration of CRISPR/Cas9 induced mutations in the genus Anastrepha. Lastly, we discuss the variables in tephritid systems that directed method development as well as recommendations for performing injections in remote containment facilities with little molecular biology capabilities. These methods and recommendations combined can serve as a guide for others to use in pursuit of developing CRISPR/Cas9 methods in tephritid systems.

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Section: Discussionmentioning

confidence: 99%

The ABCs of CRISPR in Tephritidae: developing methods for inducing heritable mutations in the genera Anastrepha, Bactrocera and Ceratitis

Sim

Kauwe

Ruano

et al. 2019

Insect Molecular Biology

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“…CRISPR/Cas9 systems developed for genome editing in D. melanogaster (Port, Chen, Lee, & Bullock, ) can be readily transferred to D. suzukii , demonstrating that the Cas9 protein and many of the promoters used to express it can function in all drosophilids. The first CRISPR/Cas9 knockouts reported in D. suzukii targeted the white and sex lethal genes and were achieved using single gRNAs that caused small deletions or two gRNAs that generated larger deletions (Kalajdzic & Schetelig, ; Li & Scott, ). Nevertheless, the low frequency of mutations in the white gene led to the assumption that batch crossing schemes, the use of plasmid DNA as the source of Cas9 and gRNAs, or the white gene itself, are responsible for the reported low targeting efficiencies (Li & Scott, ).…”

Section: Novel Biotechnology‐based Pest Control Approaches—molecular mentioning

confidence: 99%

“…The first CRISPR/Cas9 knockouts reported in D. suzukii targeted the white and sex lethal genes and were achieved using single gRNAs that caused small deletions or two gRNAs that generated larger deletions (Kalajdzic & Schetelig, 2017;Li & Scott, 2016). Nevertheless, the low frequency of mutations in the white gene led to the assumption that batch crossing schemes, the use of plasmid DNA as the source of Cas9 and gRNAs, or the white gene itself, are responsible for the reported low targeting efficiencies (Li & Scott, 2016).…”

Section: Genome Editingmentioning

confidence: 99%

Environmentally sustainable pest control options forDrosophila suzukii

Schetelig

Lee

Otto

et al. 2017

J Applied Entomology

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The spotted-wing drosophila or cherry vinegar fly (Drosophila suzukii) is native to Asia but has invaded other continents since 2008 and has spread throughout Europe. The females have a serrated ovipositor allowing them to penetrate the skins of intact ripening fruits to deposit their eggs, and the developing larvae rapidly destroy the fruits close to harvest. Drosophila suzukii has a rapid life cycle and the larvae develop well beneath the fruit surface. This means that the use of pesticides is problematic and often not effective, first due to their restricted use close to harvest to protect consumers, and second because the larvae are deep enough inside the fruit to avoid contact.There are currently no effective and environmentally sustainable pest control methods for this species, resulting in extensive damage to fruit crops. Here, we review the current status of D. suzukii as a fruit crop pest and discuss the feasibility of current pesticide-free control methods. We also consider the potential of new technologies as a basis for the urgently needed specific and long-term control of this species. K E Y W O R D Scherry vinegar fly, integrated pest management, invasive insects, RNA interference, sterile insect technique

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“…Understanding the biology and the population dynamics of D. suzukii benefits from the production and mining of genomic and transcriptomic data, as well as manipulative genetic technologies including functional transgenesis and genome editing (Kalajdzic & Schetelig, 2017;Karageorgi et al, 2017;J. Li & Handler, 2017).…”

Section: Introductionmentioning

confidence: 99%

Near-chromosome level genome assembly of the fruit pestDrosophila suzukiiusing long-read sequencing

Paris

Boyer

Jaenichen

et al. 2020

Preprint

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Over the past decade, the spotted wing Drosophila, Drosophila suzukii, has invaded Europe and America and has become a major agricultural pest in these areas, thereby prompting intense research activities to better understand its biology. Two draft genome assemblies based on short-read sequencing were released in 2013 for this species. Although valuable, these resources contain pervasive assembly errors and are highly fragmented, two features limiting their values. Our purpose here was to improve the assembly of the D. suzukii genome. For this, we generated PacBio long-read sequencing data at 160X sequence coverage and assembled a novel, contiguous D. suzukii genome. We obtained a high-quality assembly of 270 Mb (with 546 contigs, a N50 of 2.6Mb, a L50 of 15, and a BUSCO score of 95%) that we called WT3-2.0. We found that despite 16 rounds of full-sib crossings the D. suzukii strain that we sequenced has maintained high levels of polymorphism in some regions of its genome (ca. 19Mb). As a consequence, the quality of the assembly of these regions was reduced. We explored possible origins of this high residual diversity, including the presence of structural variants and a possible heterogeneous admixture pattern of North American and Asian ancestry. Overall, our WT3-2.0 assembly provides a higher quality genomic resource compared to the previous one in terms of general assembly statistics, sequence quality and gene annotation. This new D. suzukii genome assembly is therefore an improved resource for high-throughput sequencing approaches, as well as manipulative genetic technologies to study D. suzukii.

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CRISPR/Cas‐mediated gene editing using purified protein in Drosophila suzukii

Cited by 32 publications

References 65 publications

The ABCs of CRISPR in Tephritidae: developing methods for inducing heritable mutations in the genera Anastrepha, Bactrocera and Ceratitis

The ABCs of CRISPR in Tephritidae: developing methods for inducing heritable mutations in the genera Anastrepha, Bactrocera and Ceratitis

Environmentally sustainable pest control options forDrosophila suzukii

Near-chromosome level genome assembly of the fruit pestDrosophila suzukiiusing long-read sequencing

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