CasPER, a method for directed evolution in genomic contexts using mutagenesis and CRISPR/Cas9

Jakočiūnas, Tadas; Pedersen, Lasse Ebdrup; Lis, Alicia Viktoria; Jensen, Michael K.; Keasling, Jay D.

doi:10.1016/j.ymben.2018.07.001

Cited by 63 publications

(41 citation statements)

References 69 publications

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“…This plasmid pool was then used to create a genome-wide gene disruption collection, in which more than 98% of target sequences were efficiently edited with an average Increased by 3-fold with a specific titer of octadocanol at 10.3 mg/g DCW Kaczmarzyk et al, 2018 Ustilago maydis CRISPR/Cas9 Itaconic acid cyp3, MEL, UA, and P ria1 ::P etef Increased by 10.2-fold with a yield at 19.4 g/L and further enhanced to 53.5 g/L under optimized medium Becker et al, 2019 frequency of 82% (Bao et al, 2018). In parallel, Jakociunas et al employed error-prone PCR to generate DNA mutant libraries as donor, and used Cas9-mediated genome integration to introduce mutations at single-or multi-loci with efficiencies reaching 98-99%, for robust directed evolution (Jakociunas et al, 2018b). Besides, large chromosomal fragment deletion methods were developed based on CRISPR/Cas9 system.…”

Section: Promotion Of Crispr/cas System For Multi-loci Editingmentioning

confidence: 99%

“…This plasmid pool was then used to create a genome-wide gene disruption collection, in which more than 98% of target sequences were efficiently edited with an average frequency of 82% ( Bao et al, 2018 ). In parallel, Jakociunas et al employed error-prone PCR to generate DNA mutant libraries as donor, and used Cas9-mediated genome integration to introduce mutations at single- or multi-loci with efficiencies reaching 98-99%, for robust directed evolution ( Jakociunas et al, 2018b ). Besides, large chromosomal fragment deletion methods were developed based on CRISPR/Cas9 system.…”

Section: Application Of Crispr/cas System In Microbial Biotechnologymentioning

confidence: 99%

“… Halperin et al (2018) proposed a new method called EvolvR that can accelerate mutagenesis up to 7,770,000-fold within a tunable window length via CRISPR-guided nickases. Jakoèiûnas et al reported a method named Cas9-mediated Protein Evolution Reaction (CasPER) for efficient mutagenesis of nucleotides by combining error-prone PCR and Cas9-mediated genome integration ( Jakociunas et al, 2018b ). Garst et al (2017) constructed CRISPR-enabled trackable genome engineering (CREATE) method, where a library of targets was built and transformed for multiplex editing in vivo , followed by screening and mutation identification.…”

Section: Application Of Crispr/cas System In Microbial Biotechnologymentioning

confidence: 99%

See 2 more Smart Citations

Development and Application of CRISPR/Cas in Microbial Biotechnology

Ding

Yang

Shi

2020

Front. Bioeng. Biotechnol.

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The clustered regularly interspaced short palindromic repeats (CRISPR)-associated (Cas) system has been rapidly developed as versatile genomic engineering tools with high efficiency, accuracy and flexibility, and has revolutionized traditional methods for applications in microbial biotechnology. Here, key points of building reliable CRISPR/Cas system for genome engineering are discussed, including the Cas protein, the guide RNA and the donor DNA. Following an overview of various CRISPR/Cas tools for genome engineering, including gene activation, gene interference, orthogonal CRISPR systems and precise single base editing, we highlighted the application of CRISPR/Cas toolbox for multiplexed engineering and high throughput screening. We then summarize recent applications of CRISPR/Cas systems in metabolic engineering toward production of chemicals and natural compounds, and end with perspectives of future advancements.

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Section: Promotion Of Crispr/cas System For Multi-loci Editingmentioning

confidence: 99%

Section: Application Of Crispr/cas System In Microbial Biotechnologymentioning

confidence: 99%

Section: Application Of Crispr/cas System In Microbial Biotechnologymentioning

confidence: 99%

See 1 more Smart Citation

Development and Application of CRISPR/Cas in Microbial Biotechnology

Ding

Yang

Shi

2020

Front. Bioeng. Biotechnol.

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“…; Jakočiūnas et al. ) could also be utilized for genetic barcoding projects. Because most current genome‐editing approaches require the manipulation of zygotes, the technology will primarily be applicable to species for which gametes can be collected, fertilized, and the resulting offspring grown to maturation in the wild or in captivity.…”

Section: Genome Editing In Organismsmentioning

confidence: 99%

Transforming ecology and conservation biology through genome editing

Phelps

Seeb

2019

Conservation Biology

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As the conservation challenges increase, new approaches are needed to help combat losses in biodiversity and slow or reverse the decline of threatened species. Genome-editing technology is changing the face of modern biology, facilitating applications that were unimaginable only a decade ago. The technology has the potential to make significant contributions to the fields of evolutionary biology, ecology, and conservation, yet the fear of unintended consequences from designer ecosystems containing engineered organisms has stifled innovation. To overcome this gap in the understanding of what genome editing is and what its capabilities are, more research is needed to translate genome-editing discoveries into tools for ecological research. Emerging and future genome-editing technologies include new clustered regularly interspaced short palindromic repeats (CRISPR) targeted sequencing and nucleic acid detection approaches as well as species genetic barcoding and somatic genome-editing technologies. These genome-editing tools have the potential to transform the environmental sciences by providing new noninvasive methods for monitoring threatened species or for enhancing critical adaptive traits. A pioneering effort by the conservation community is required to apply these technologies to real-world conservation problems.

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“…Together with large pooled single-guide RNA (sgRNA) libraries, CRISPR-Cas proteins or fusion proteins coupled with the effectors can serve as screening tools [121], such as screening cancer-related genomic ribonucleotides [122], identifying fetal hemoglobin regulator [123] and so on. The CRISPR-Cas system can also serve as a nucleic acid mutation toolkit [124]. Taking advantages of the CRISPR system, researchers can create recording tools [125] and evolutional tools with nCas9 (D10A) combined with the fidelity-reduced Escherichia coli DNA polymerase I [126].…”

Section: Development Of Crispr Systemsmentioning

confidence: 99%

Blossom of CRISPR technologies and applications in disease treatment

Liu

Wang

Luo

2018

Synthetic and Systems Biotechnology

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Since 2013, the CRISPR-based bacterial antiviral defense systems have revolutionized the genome editing field. In addition to genome editing, CRISPR has been developed as a variety of tools for gene expression regulations, live cell chromatin imaging, base editing, epigenome editing, and nucleic acid detection. Moreover, in the context of further boosting the usability and feasibility of CRISPR systems, novel CRISPR systems and engineered CRISPR protein mutants have been explored and studied actively. With the flourish of CRISPR technologies, they have been applied in disease treatment recently, as in gene therapy, cell therapy, immunotherapy, and antimicrobial therapy. Here we present the developments of CRISPR technologies and describe the applications of these CRISPR-based technologies in disease treatment.

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CasPER, a method for directed evolution in genomic contexts using mutagenesis and CRISPR/Cas9

Cited by 63 publications

References 69 publications

Development and Application of CRISPR/Cas in Microbial Biotechnology

Development and Application of CRISPR/Cas in Microbial Biotechnology

Transforming ecology and conservation biology through genome editing

Blossom of CRISPR technologies and applications in disease treatment

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