Novel CRISPR/Cas applications in plants: from prime editing to chromosome engineering DOI Creative Commons
Teng‐Kuei Huang, Holger Puchta

Transgenic Research, Journal Year: 2021, Volume and Issue: 30(4), P. 529 - 549

Published: March 1, 2021

Abstract In the last years, tremendous progress has been made in development of CRISPR/Cas-mediated genome editing tools. A number natural CRISPR/Cas nuclease variants have characterized. Engineered Cas proteins developed to minimize PAM restrictions, off-side effects and temperature sensitivity. Both kinds enzymes have, by now, applied widely efficiently many plant species generate either single or multiple mutations at desired loci multiplexing. addition DSB-induced mutagenesis, specifically designed systems allow more precise gene editing, resulting not only random but also predefined changes. Applications plants include targeting homologous recombination, base and, recently, prime editing. We will evaluate these different technologies for their prospects practical applicability plants. addition, we discuss a novel application Cas9 plants, enabling induction heritable chromosomal rearrangements, such as inversions translocations. This technique make it possible change genetic linkages programmed way add another level engineering toolbox breeding. Also, strategies tissue culture free were developed, which might be helpful overcome transformation bottlenecks crops. All all, recent advances technology help agriculture address challenges twenty-first century related global warming, pollution food shortage.

Language: Английский

Genome engineering for crop improvement and future agriculture DOI Creative Commons
Caixia Gao

Cell, Journal Year: 2021, Volume and Issue: 184(6), P. 1621 - 1635

Published: Feb. 15, 2021

Language: Английский

Citations

681
Applications of CRISPR–Cas in agriculture and plant biotechnology DOI
Haocheng Zhu, Chao Li, Caixia Gao

et al.

Nature Reviews Molecular Cell Biology, Journal Year: 2020, Volume and Issue: 21(11), P. 661 - 677

Published: Sept. 24, 2020

Language: Английский

Citations

652
Prime editing for precise and highly versatile genome manipulation DOI
Peter J. Chen, David R. Liu

Nature Reviews Genetics, Journal Year: 2022, Volume and Issue: 24(3), P. 161 - 177

Published: Nov. 7, 2022

Language: Английский

Citations

324
The CRISPR-Cas toolbox and gene editing technologies DOI Creative Commons

Guanwen Liu,

Qiupeng Lin, Shuai Jin

et al.

Molecular Cell, Journal Year: 2021, Volume and Issue: 82(2), P. 333 - 347

Published: Dec. 29, 2021

Language: Английский

Citations

264
Precise plant genome editing using base editors and prime editors DOI
Kutubuddin A. Molla, Simon Sretenovic, K. C. Bansal

et al.

Nature Plants, Journal Year: 2021, Volume and Issue: 7(9), P. 1166 - 1187

Published: Sept. 13, 2021

Language: Английский

Citations

256
CRISPR/Cas: A powerful tool for gene function study and crop improvement DOI Creative Commons
Dangquan Zhang, Zhiyong Zhang, Turgay Ünver

et al.

Journal of Advanced Research, Journal Year: 2020, Volume and Issue: 29, P. 207 - 221

Published: Oct. 21, 2020

It is a long-standing goal of scientists and breeders to precisely control gene for studying its function as well improving crop yield, quality, tolerance various environmental stresses. The discovery modification CRISPR/Cas system, nature-occurred editing tool, opens an era precision breeding.In this review, we first introduce the brief history followed mechanism application system on study improvement. Currently, genome has been becoming mature cutting-edge biotechnological tool improvement that already used in many different traits crops, including pathogen resistance, abiotic tolerance, plant development morphology even secondary metabolism fiber development. Finally, point out major issues associating with future research directions.Key Scientific Concepts Review: CRISPR/Cas9 robust powerful targeting individual DNA RNA sequence genome. can be target knockin, knockout replacement monitoring regulating expression at epigenome levels by binding specific sequence. Agrobacterium-mediated method still efficient delivering regents into targeted cells. However, other delivery methods, such virus-mediated method, have developed enhanced potentials CRISPR/Cas9-based PAM requirement offers CRISPR/Cas9-targted genetic loci also limits CRISPR/Cas9. Discovering new Cas proteins modifying current enzymes play important role editing. Developing better methods eliminating off-target effects, finding key/master genes controlling growth two directions

Language: Английский

Citations

253
High-efficiency prime editing with optimized, paired pegRNAs in plants DOI
Qiupeng Lin, Shuai Jin, Yuan Zong

et al.

Nature Biotechnology, Journal Year: 2021, Volume and Issue: 39(8), P. 923 - 927

Published: March 25, 2021

Language: Английский

Citations

247
Prime editing efficiently generates W542L and S621I double mutations in two ALS genes in maize DOI Creative Commons
Yuanyuan Jiang, Yiping Chai, Minhui Lu

et al.

Genome biology, Journal Year: 2020, Volume and Issue: 21(1)

Published: Oct. 6, 2020

Prime editing is a novel and universal CRISPR/Cas-derived precision genome-editing technology that has been recently developed. However, low efficiency of prime shown in transgenic rice lines. We hypothesize enhancing pegRNA expression could improve prime-editing efficiency. In this report, we describe two strategies for expression. construct vector harboring variants W542L S621I double mutations ZmALS1 ZmALS2. Compared with previous reports rice, achieve much higher maize. Our results are inspiring provide direction the optimization plant editors.

Language: Английский

Citations

210
Prime editing for functional repair in patient-derived disease models DOI Creative Commons
Imre F. Schene,

Indi P. Joore,

Rurika Oka

et al.

Nature Communications, Journal Year: 2020, Volume and Issue: 11(1)

Published: Oct. 23, 2020

Abstract Prime editing is a recent genome technology using fusion proteins of Cas9-nickase and reverse transcriptase, that holds promise to correct the vast majority genetic defects. Here, we develop prime for primary adult stem cells grown in organoid culture models. First, generate precise in-frame deletions gene encoding β‐catenin ( CTNNB1 ) result proliferation independent Wnt-stimuli, mimicking mechanism development liver cancer. Moreover, functionally recovers disease-causing mutations intestinal organoids from patients with DGAT1-deficiency patient Wilson disease ATP7B ). as efficient 3D 2D cell lines offers greater precision than Cas9-mediated homology directed repair (HDR). Base remains more reliable but restricted subgroup pathogenic mutations. Whole-genome sequencing four prime-edited clonal reveals absence genome-wide off-target effects underscoring therapeutic potential this versatile strategy.

Language: Английский

Citations

187
Precision genome engineering in rice using prime editing system DOI Creative Commons
Kai Hua, Yuwei Jiang, Xiaoping Tao

et al.

Plant Biotechnology Journal, Journal Year: 2020, Volume and Issue: 18(11), P. 2167 - 2169

Published: May 6, 2020

Dear editor, Making precise changes in the genomes of organisms is challenging for most genome editing tools. Recently, a search-and-replace method, also known as prime editing, was developed that can introduce user-defined sequence into target site without requiring double-stranded breaks (DSBs) or repair templates (Anzalone et al., 2019). The editor contains Moloney murine leukaemia virus reverse transcriptase (M-MLV RT) fused to C terminus SpCas9 (H840A) nickase This fusion protein guided by guide RNA (pegRNA) site. In addition specifying site, pegRNA primer binding (PBS) which complementary PAM-containing strand and template transcription (i.e. RT sequence). genetic information be introduced encoded sequence. editors all 12 base-to-base conversions, small indels their combinations. Therefore, they hold great promise gene therapy well precision breeding crops. Here, we report application engineering rice plants. We synthesized an engineered M-MLV (D200N/L603W/T306K/ W313F/T330P) 2019) used it construct Sp-PE2 Sp-PE3 expression (Figure 1a). Compared Sp-PE2, express additional nick sgRNA To test whether are functional plant cells, transgenic reporter monitor activity calli. constructed cassette containing inactive EGFP driven CaMV 35S promoter 1b) inserted Sp-PE3. Both Y67 G68, two essential chromophore residues EGFP, were changed stop codons 1b). Only base conversions (T-G G-C) restore wild-type sequence, whereas other forms cannot. designed with 13 nt PBS targeting 1c) loaded targets 47 away from pegRNA-induced nick. T-DNA vectors calli through Agrobacterium-mediated transformation. After integration genome, expressed would edit under guidance pegRNA. weeks selection, found more than fifty per cent transformed showed GFP signals (Figures 1d,e). Sp-PE3, on opposite strand, did not show increased ratio positive one month randomly selected three hygromycin-resistant signal genotyping indeed harboured 1f,g). contrast, any mutation region then regenerated (with signals) obtained 32 35 lines respectively. genotyping, frequencies comparable between (15.6% 17.1%) 1l). One line edited contained only restored 1h), had no at but nicking rest both original endogenous genes, first chose acetolactate synthase (ALS) target. A 16 84 downstream ALS S627N mutation, makes plants resistant imidazolinone herbicides 1i). 4 out 44 (9.1%) desired G-A transition detected 1j,l). Among four lines, heterozygous chimeric. C42F ABERRANT PANICLE ORGANIZATION 1 (APO1). However, this These results indicate generate efficiency varies different sites. slightly higher may due copy number genome. tested ability deletions insertions could events even indel mutations targeted sites SaCas9 multiple-turnover enzyme releases cleaved DNA products faster does (Yourik hypothesized replacing (N580A) increase because facilitate carry transcription. Sa-PE3 1c). 78 very week after selection Genotyping indicated fraction cells have 1k). Consequently, find event stable Next, Taken together, these suggest has lower compared As structure scaffold different, SaCas9-PE3 require further optimization improve efficiency. summary, tool precisely rice. During preparation review our manuscript, several systems wheat reported (Li 2020; Lin Tang Xu 2020a; 2020b). colleagues presented data protoplasts obtain (Tang 2020), 1-3 genes relatively low efficiencies studies. It possible system improved using transcriptases (Stamos 2017) modifying transgene make powerful tools molecular work supported National Natural Science Foundation China (No.U19A2022) Chinese Academy Sciences. K.H. partly Postdoctoral (2019M661649) Shanghai Super Incentive Program. authors declare conflict interests. K.H., W.J., X.T. performed experiments analysed data. wrote manuscript. J.-K. Z. supervised project

Language: Английский

Citations

148