Precise insertion and guided editing of higher plant genomes using Cpf1 CRISPR nucleases

Begemann, Matthew B.; Gray, Benjamin N.; January, Emma; Gordon, Gina C.; He, Yanni; Liu, Haijun; Wu, Xingrong; Brutnell, Thomas P.; Mockler, Todd C.; Oufattole, Mohammed

doi:10.1101/109983

Cited by 18 publications

(27 citation statements)

References 26 publications

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“…BV3L6 (AsCas12a) and Lachnospiraceae bacterium ND2006 as active in human cells [64]. In rice, using FnCas12a and LbCas12a targeted insertions via HR were accomplished and, at least for FnCas12a, with higher rates than SpCas9-based experiments [76]. However, more recent experiments revealed that FnCas12a possesses robust DNA cleavage activity in human cells as well with frequencies comparable to those of the other orthologues [67].…”

Section: Using Cas12a (Formerly Named Cpf1) In Plantsmentioning

confidence: 99%

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

2018

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Currently, biology is revolutionized by ever growing applications of the CRISPR/Cas system. As discussed in this Review, new avenues have opened up for plant research and breeding by the use of the sequence-specific DNases Cas9 and Cas12 (formerly named Cpf1) and, more recently, the RNase Cas13 (formerly named C2c2). Although double strand break-induced gene editing based on error-prone nonhomologous end joining has been applied to obtain new traits, such as powdery mildew resistance in wheat or improved pathogen resistance and increased yield in tomato, improved technologies based on CRISPR/Cas for programmed change in plant genomes via homologous recombination have recently been developed. Cas9- and Cas12- mediated DNA binding is used to develop tools for many useful applications, such as transcriptional regulation or fluorescence-based imaging of specific chromosomal loci in plant genomes. Cas13 has recently been applied to degrade mRNAs and combat viral RNA replication. By the possibility to address multiple sequences with different guide RNAs and by the simultaneous use of different Cas proteins in a single cell, we should soon be able to achieve complex changes of plant metabolism in a controlled way.

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Section: Using Cas12a (Formerly Named Cpf1) In Plantsmentioning

confidence: 99%

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

2018

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“…Genome editing is a powerful tool for increasing plant yields, improving food quality, enhancing crop disease resistance and developing new cultivars to meet market needs (Begemann et al ., ). At present, several strategies are being exploited to edit plant genomes, including CRISPR‐Cas, meganucleases, TALENs and zinc finger nucleases (Martín‐Pizarro and Posé, ).…”

Section: Introductionmentioning

confidence: 97%

“…To date, Acidaminococcus sp. BV3L6 Cas12a (AsCas12a), Francisella novicida Cas12a (FnCas12a) and Lachnospiraceae bacterium ND2006 Cas12a (LbCas12a) have been used to edit the genomes of crop and model plants, including green alga, rice, soybeans and tobacco (Begemann et al ., ; Endo et al ., ; Ferenczi et al ., ; Hu et al ., ; Kim et al ., ; Tang et al ., ; Wang et al ., ,b,c; Xu et al ., ; Yin et al ., ), but not citrus. In addition, the performances of the three Cas12a homologs are different.…”

Section: Introductionmentioning

confidence: 98%

CRISPR‐LbCas12a‐mediated modification of citrus

Jia

Orbović

Wang

2019

Plant Biotechnology Journal

150

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Summary Recently, CRISPR‐Cas12a (Cpf1) from Prevotella and Francisella was engineered to modify plant genomes. In this report, we employed CRISPR‐LbCas12a (LbCpf1), which is derived from Lachnospiraceae bacterium ND2006, to edit a citrus genome for the first time. First, LbCas12a was used to modify the CsPDS gene successfully in Duncan grapefruit via Xcc‐facilitated agroinfiltration. Next, LbCas12a driven by either the 35S or Yao promoter was used to edit the PthA4 effector binding elements in the promoter (EBEPthA4‐CsLOBP) of CsLOB1. A single crRNA was selected to target a conserved region of both Type I and Type II CsLOBPs, since the protospacer adjacent motif of LbCas12a (TTTV) allows crRNA to act on the conserved region of these two types of CsLOBP. CsLOB1 is the canker susceptibility gene, and it is induced by the corresponding pathogenicity factor PthA4 in Xanthomonas citri by binding to EBEPthA4‐CsLOBP. A total of seven 35S‐LbCas12a‐transformed Duncan plants were generated, and they were designated as #D35s1 to #D35s7, and ten Yao‐LbCas12a‐transformed Duncan plants were created and designated as #Dyao1 to #Dyao10. LbCas12a‐directed EBEPthA4‐CsLOBP modifications were observed in three 35S‐LbCas12a‐transformed Duncan plants (#D35s1, #D35s4 and #D35s7). However, no LbCas12a‐mediated indels were observed in the Yao‐LbCas12a‐transformed plants. Notably, transgenic line #D35s4, which contains the highest mutation rate, alleviates XccΔpthA4:dCsLOB1.4 infection. Finally, no potential off‐targets were observed. Therefore, CRISPR‐LbCas12a can readily be used as a powerful tool for citrus genome editing.

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“…Numerous GEEN reagents-engineered meganucleases, zinc finger nucleases (ZFNs), transcriptional activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats and associated Cas9 endonuclease (CRISPR/Cas9)-have been developed and extensively reviewed for their efficiency, specificity, and application to various plant systems [15,16]. The emergence of the CRISPR/ Cas9 reagent system with its relative ease of use, its modification by use of single-guide RNA (sgRNA) [17], and its improvement through redesign of the Cas9 nuclease [18] or adoption of alternative nuclease systems, such as Cpf1 [19], has led to the rapid and widespread use of genome editing as a preferred method to target highly specific changes to plant genomes.…”

Section: Diversity Of Tools and Outcomes For Genome Editingmentioning

confidence: 99%

Risk associated with off-target plant genome editing and methods for its limitation

Zhao

Wolt

2017

Emerging Topics in Life Sciences

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Assessment for potential adverse effects of plant genome editing logically focuses on the specific characteristics of the derived phenotype and its release environment. Genome-edited crops, depending on the editing objective, can be classified as either indistinguishable from crops developed through conventional plant breeding or as crops which are transgenic. Therefore, existing regulatory regimes and risk assessment procedures accommodate genome-edited crops. The ability for regulators and the public to accept a product focus in the evaluation of genome-edited crops will depend on research which clarifies the precision of the genome-editing process and evaluates unanticipated off-target edits from the process. Interpretation of genome-wide effects of genome editing should adhere to existing frameworks for comparative risk assessment where the nature and degree of effects are considered relative to a baseline of genome-wide mutations as found in crop varieties developed through conventional breeding methods. Research addressing current uncertainties regarding unintended changes from plant genome editing, and adopting procedures that clearly avoid the potential for gene drive initiation, will help to clarify anticipated public and regulatory questions regarding risk of crops derived through genome editing.

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Precise insertion and guided editing of higher plant genomes using Cpf1 CRISPR nucleases

Cited by 18 publications

References 26 publications

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

CRISPR‐LbCas12a‐mediated modification of citrus

Risk associated with off-target plant genome editing and methods for its limitation

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