Cpf1-Database: web-based genome-wide guide RNA library design for gene knockout screens using CRISPR-Cpf1

Park, Jeongbin; Bae, Sangsu

doi:10.1093/bioinformatics/btx695

Cited by 23 publications

(13 citation statements)

References 18 publications

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“…In this study, most of the spacer used were designed via http://www.rgenome.net (Park & Bae, ; Table S2). To test the efficiency of the iCOPE method, a four‐fluorescent protein or chromoprotein (mRFP, eYFP, amilCP, and cjBlue) cassette was introduced into pUC19 to generate pUC19.S4.…”

Section: Resultsmentioning

confidence: 99%

Improved CRISPR‐Cas12a‐assisted one‐pot DNA editing method enables seamless DNA editing

Wang

Liu

et al. 2019

Biotech & Bioengineering

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As the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas12a (previously known as Cpf1) system cleaves double-stranded DNA and produces a sticky end, it could serve as a useful tool for DNA assembly/editing. To broaden its application, a variety of engineered FnCas12a proteins are generated with expanded protospacer adjacent motif (PAM) requirements. Two variants (FnCas12a-EP15 and EP16) increased the targeting range of FnCas12a by approximately fourfold. They can efficiently recognize a broad range of PAM sequences including YN (Y = C or T), TAC and CAA. Meanwhile, based on our demonstration that FnCas12a is active from 16 to 60°C, we developed an "improved CRISPR-Cas12a-assisted one-pot DNA editing" (iCOPE) method to facilitate DNA editing by combining the crRNA transcription, digestion, and ligation in one pot. By applying iCOPE, the editing efficiency reached 72-100% for two DNA fragment assemblies, and for the 21 kb large DNA construct modification, the editing efficiency can reach 100%. Thanks to the advantages of Cas12a, iCOPE with only one digestion enzyme could replace current a variety of restriction enzymes to perform the cloning in one pot with almost no sequence constraints. Taken together, this study offers an expanded DNA targeting scope of CRISPR systems and could serve as an efficient seamless one-pot DNA editing tool. K E Y W O R D SCas12a variants, DNA editing, FnCas12a, one pot, synthetic biology

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Section: Resultsmentioning

confidence: 99%

Improved CRISPR‐Cas12a‐assisted one‐pot DNA editing method enables seamless DNA editing

Wang

Liu

et al. 2019

Biotech & Bioengineering

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“…Cpf1-Database is another web-based gRNA design tool, which includes all the available pre-searched targets of Cpf1 with 5′-TTTN-3′ PAM sequences in all coding sequence (CDS) regions within the whole genome of 12 selected organisms. Cpf1-Database provides a rapid and simple way of designing gRNAs for thousands of genes and facilitates the genome-wide screening experiment via usage of Cpf1 [37]. Cpf1-Database is updated frequently by following the recent gene/transcript/CDS annotation information and alternative PAMs of Cpf1 (Table 1).…”

Section: Guide Rna Designmentioning

confidence: 99%

CRISPR Cpf1 proteins: structure, function and implications for genome editing

et al. 2019

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CRISPR and CRISPR-associated (Cas) protein, as components of microbial adaptive immune system, allows biologists to edit genomic DNA in a precise and specific way. CRISPR-Cas systems are classified into two main classes and six types. Cpf1 is a putative type V (class II) CRISPR effector, which can be programmed with a CRISPR RNA to bind and cleave complementary DNA targets. Cpf1 has recently emerged as an alternative for Cas9, due to its distinct features such as the ability to target T-rich motifs, no need for trans-activating crRNA, inducing a staggered double-strand break and potential for both RNA processing and DNA nuclease activity. In this review, we attempt to discuss the evolutionary origins, basic architectures, and molecular mechanisms of Cpf1 family proteins, as well as crRNA designing and delivery strategies. We will also describe the novel Cpf1 variants, which have broadened the versatility and feasibility of this system in genome editing, transcription regulation, epigenetic modulation, and base editing. Finally, we will be reviewing the recent studies on utilization of Cpf1as a molecular tool for genome editing.

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“…The PAM sequence may vary according to its origin of ortholog. The Cpf1-database 1 is an online tool that offers a simple and easy way to find a potential target within the genome and design gRNA, and it can recognize AsCpf1 and LbCpf1 nucleases through DNA recognition sequences (Park and Bae, 2017).…”

Section: Introductionmentioning

confidence: 99%

The Rise of the CRISPR/Cpf1 System for Efficient Genome Editing in Plants

et al. 2020

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Cpf1, an endonuclease of the class 2 CRISPR family, fills the gaps that were previously faced in the world of genome engineering tools, which include the TALEN, ZFN, and CRISPR/Cas9. Other simultaneously discovered nucleases were not able to carry out reengineering at the same region due to the loss of a target site after first-time engineering. Cpf1 acts as a dual nuclease, functioning as an endoribonuclease to process crRNA and endodeoxyribonuclease to cleave target sequences and generate double-stranded breaks. Additionally, Cpf1 allows for multiplexed genome editing, as a single crRNA array transcript can target multiple loci in the genome. The CRISPR/Cpf1 system enables gene deletion, insertion, base editing, and locus tagging in monocot as well as in dicot plants with fewer off-target effects. This tool has been efficiently demonstrated into tobacco, rice, soybean, wheat, etc. This review covers the development and applications of Cpf1 mediated genome editing technology in plants.

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Cpf1-Database: web-based genome-wide guide RNA library design for gene knockout screens using CRISPR-Cpf1

Abstract: sangsubae@hanyang.ac.kr.

Cited by 23 publications

References 18 publications

Improved CRISPR‐Cas12a‐assisted one‐pot DNA editing method enables seamless DNA editing

Improved CRISPR‐Cas12a‐assisted one‐pot DNA editing method enables seamless DNA editing

CRISPR Cpf1 proteins: structure, function and implications for genome editing

The Rise of the CRISPR/Cpf1 System for Efficient Genome Editing in Plants

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