Research Progress on Off-target Effects and Detection Techniques in CRISPR/Cas9 Systems

doi:10.13560/j.cnki.biotech.bull.1985.2019-0871

Abstract

Abstract: Clustered regularly interspaced short palindromic repeats（CRISPR）and its associated protein Cas9 have become as a powerful tool for precise genome editing. The CRISPR/Cas9 system,derived from the immune mechanism of bacteria and archaea,is simple in structure and easy to design. Different gene site can be quickly edited by simply changing a small sequence on the sgRNA. Compared with the zinc finger nucleases and transcription activator-like effector nucleases,the gene editing efficiency by CRISPR/Cas9 system is higher and the specificity is stronger. However,the inevitable off-target event in the application process severely limits the further development of the CRISPR/Cas9 system. Therefore,this article reviews the off-target type,influencing factors,reduction strategies and off-target detection technologies in applying the CRISPR/Cas9 system.

Key words: CRISPR/Cas9, off-target effect, influencing factors, reduction strategy, detection technology

ZHANG Chen, LEI Zhan, LI Kai, SHANG Ying, XU Wen-tao. Research Progress on Off-target Effects and Detection Techniques in CRISPR/Cas9 Systems[J]. Biotechnology Bulletin, 2020, 36(3): 78-87.

References

[1] Ishino Y, Shinagawa H, Makino K, et al.Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product[J]. Journal of Bacteriology, 1987, 169(12):5429-5433.
[2] Jansen R, Embden JDAV, Gaastra W, et al.Identification of genes that are associated with DNA repeats in prokaryotes[J]. Molecular Microbiology, 2010, 43(6):1565-1575.
[3] Barrangou R, Fremaux C, Deveau H, et al.CRISPR provides acquired resistance against viruses in prokaryotes[J]. Science, 2007, 315(5819):1709-1712.
[4] Le C, Ran FA, Cox D, et al.Multiplex genome engineering using CRISPR/Cas systems[J]. Science, 2015, 1239(11):197.
[5] Jiang W, Bikard D, Cox D, et al.RNA-guided editing of bacterial genomes using CRISPR-Cas systems[J]. Nat Biotechnol, 2013, 31(3):233-239.
[6] Horvath P, Barrangou R.CRISPR/Cas, the immune system of bacteria and archaea[J]. Science, 2010, 327(5962):167-170.
[7] Koonin EV, Makarova KS, Zhang F.Diversity, classification and evolution of CRISPR-Cas systems[J]. Curr Opin Microbiol, 2017, 37:67-78.
[8] Ishino Y, Krupovic M, Forterre P.History of CRISPR-Cas from encounter with a mysterious repeated sequence to genome editing technology[J]. Journal of Bacteriology, 2018, 200(7):1-16.
[9] Cong L, Ran FA, Cox D, et al.Multiplex genome engineering using CRISPR/Cas systems[J]. Science, 2013, 339(6121):819-823.
[10] Javed MR, Sadaf M, Ahmed T, et al.CRISPR-Cas system:history and prospects as a genome editing tool in microorganisms[J]. Curr Microbiol, 2018, 75(12):1675-1683.
[11] Gasiunas G, Siksnys V.RNA-dependent DNA endonuclease Cas9 of the CRISPR system:holy grail of genome editing?[J]. Trends in Microbiology, 2013, 21(11):562-567.
[12] Yin H, Xue W, Anderson DG.CRISPR-Cas:a tool for cancer research and therapeutics[J]. Nature Reviews Clinical Oncology, 2019, 16(5):281-295.
[13] Shrock E, Guell M.CRISPR in animals and animal models[J]. Progress in Molecular Biology and Translational Science, 2017, 152:95-114.
[14] Yin K, Gao C, Qiu JL.Progress and prospects in plant genome editing[J]. Nature Plants, 2017, 3:17107.
[15] Bolukbasi MF, Gupta A, Wolfe SA.Creating and evaluating accurate CRISPR-Cas9 scalpels for genomic surgery[J]. Nat Methods, 2016, 13(1):41-50.
[16] Fu Y, Foden JA, Khayter C, et al.High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells[J]. Nat Biotechnol, 2013, 31(9):822-826.
[17] Cradick TJ, Fine EJ, Antico CJ, et al.CRISPR/Cas9 systems targeting beta-globin and CCR5 genes have substantial off-target activity[J]. Nucleic Acids Res, 2013, 41(20):9584-9592.
[18] Lin Y, Cradick TJ, Brown MT, et al.CRISPR/Cas9 systems have off-target activity with insertions or deletions between target DNA and guide RNA sequences[J]. Nucleic Acids Research, 2014, 42(11):7473-7485.
[19] Zhang XH, Tee LY, Wang XG, et al.Off-target effects in CRISPR/Cas9-mediated genome engineering[J]. Molecular Therapy-Nucleic Acids, 2015, 4(11):e264.
[20] Doench JG, Fusi N, Sullender M, et al.Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9[J]. Nat Biotechnol, 2016, 34(2):184.
[21] Hruscha A, Krawitz P, Rechenberg A, et al.Efficient CRISPR/Cas9 genome editing with low off-target effects in zebrafish[J]. Development, 2013, 140(24):4982-4987.
[22] Zhu LJ.Overview of guide RNA design tools for CRISPR-Cas9 genome editing technology[J]. Frontiers in Biology, 2015, 10(4):289-296.
[23] Gilbert LA, Horlbeck MA, Adamson B, et al.Genome-scale CRISPR-mediated control of gene repression and activation[J]. Cell, 2014, 159(3):647-661.
[24] Cho SW, Kim S, Kim Y, et al.Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases and nickases[J]. Genome Research, 2014, 24(1):132-141.
[25] Hsu PD, Scott DA, Weinstein JA, et al.DNA targeting specificity of RNA-guided Cas9 nucleases[J]. Nat Biotechnol, 2013, 31(9):827.
[26] Yin H, Song CQ, Suresh S, et al.Partial DNA-guided Cas9 enables genome editing with reduced off-target activity[J]. Nature Chemical Biology, 2018, 14(3):311-316.
[27] Frock RL, Hu J, Meyers RM, et al.Genome-wide detection of DNA double-stranded breaks induced by engineered nucleases[J]. Nat Biotechnol, 2015, 33(2):179.
[28] Guilinger JP, Thompson DB, Liu DR.Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification[J]. Nat Biotechnol, 2014, 32(6):577-582.
[29] Zhu X, Clarke R, Puppala AK, et al.Cryo-EM structures reveal coordinated domain motions that govern DNA cleavage by Cas9[J]. Nature Structural & Molecular Biology, 2019, 26(8):679-685.
[30] Kleinstiver BP, Pattanayak V, Prew MS, et al.High-fidelity CRISPR-Cas9 nucleases with no detectable genome-wide off-target effects[J]. Nature, 2016, 529(7587):490-495.
[31] Slaymaker IM, Gao L, Zetsche B, et al.Rationally engineered Cas9 nucleases with improved specificity[J]. Science, 2016, 351(6268):84-88.
[32] Chen JS, Dagdas YS, Kleinstiver BP, et al.Enhanced proofreading governs CRISPR-Cas9 targeting accuracy[J]. Nature, 2017, 550(7676):407-410.
[33] Casini A, Olivieri M, Petris G, et al.A highly specific SpCas9 variant is identified by in vivo screening in yeast[J]. Nat Biotechnol, 2018, 36(3):265-271.
[34] Hu JH, Miller SM, Geurts MH, et al.Evolved Cas9 variants with broad PAM compatibility and high DNA specificity[J]. Nature, 2018, 556(7699):57-63.
[35] Lee JK, Jeong E, Lee J, et al.Directed evolution of CRISPR-Cas9 to increase its specificity[J]. Nature Communications, 2018, 9
(1):3048.
[36] Vakulskas CA, Dever DP, Rettig GR, et al.A high-fidelity Cas9 mutant delivered as a ribonucleoprotein complex enables efficient gene editing in human hematopoietic stem and progenitor cells[J]. Nature Medicine, 2018, 24(8):1216-1224.
[37] Nishimasu H, Cong L, Yan WX, et al.Crystal structure of Staphylococcus aureus Cas9[J]. Cell, 2015, 162(5):1113-1126.
[38] Müller M, Lee CM, Gasiunas G, et al.Streptococcus thermophilus CRISPR-Cas9 systems enable specific editing of the human genome[J]. Molecular Therapy, 2016, 24(3):636-644.
[39] Xu K, Ren C, Liu Z, et al.Efficient genome engineering in eukaryotes using Cas9 from Streptococcus thermophilus[J]. Cellular and Molecular Life Sciences, 2015, 72(2):383-399.
[40] Lee CM, Cradick TJ, Bao G.The neisseria meningitidis CRISPR-Cas9 system enables specific genome editing in mammalian cells[J]. Molecular Therapy, 2016, 24(3):645-654.
[41] Murovec J, Pirc Z, Yang B.New variants of CRISPR RNA-guided genome editing enzymes[J]. Plant Biotechnology Journal, 2017, 15(8):917-926.
[42] Kim E, Koo T, Park SW, et al.In vivo genome editing with a small Cas9 orthologue derived from Campylobacter jejuni[J]. Nature Communications, 2017, 8:14500.
[43] Knott GJ, Thornton BW, Lobba MJ, et al.Broad-spectrum enzymatic inhibition of CRISPR-Cas12a[J]. Nature Structural & Molecular Biology, 2019, 26(4):315-321.
[44] Watters KE, Fellmann C, Bai HB, et al.Systematic discovery
of natural CRISPR-Cas12a inhibitors[J]. Science, 2018, 362(6411):236-239.
[45] Strecker J, Jones S, Koopal B, et al.Engineering of CRISPR-Cas12b for human genome editing[J]. Nature Communications, 2019, 10(1):212.
[46] Ran FA, Cong L, Yan WX, et al.In vivo genome editing using Staphylococcus aureus Cas9[J]. Nature, 2015, 520(7546):186-191.
[47] Kleinstiver BP, Prew MS, Tsai SQ, et al.Broadening the targeting range of Staphylococcus aureus CRISPR-Cas9 by modifying PAM recognition[J]. Nat Biotechnol, 2015, 33(12):1293-1298.
[48] Pawluk A, Amrani N, Zhang Y, et al.Naturally occurring off-switches for CRISPR-Cas9[J]. Cell, 2016, 167(7):1829-1838.
[49] Chen F, Ding X, Feng Y, et al.Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting[J]. Nature Communications, 2017, 8:14958.
[50] Gao Z, Herrera Carrillo E, Berkhout B.A single H1 promoter can drive both guide RNA and endonuclease expression in the CRISPR-Cas9 system[J]. Molecular Therapy Nucleic Acids, 2019, 14:32-40.
[51] Hendel A, Fine EJ, Bao G, et al.Quantifying on- and off-target genome editing[J]. Trends Biotechnol, 2015, 33(2):132-140.
[52] Cho SW, Kim S, Kim Y, et al.Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases and nickases[J]. Genome Res, 2014, 24(1):132-141.
[53] Bae S, Park J, Kim JS.Cas-OFFinder:a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases[J]. Bioinformatics, 2014, 30(10):1473-1475.
[54] Tsai SQ, Joung JK.Defining and improving the genome-wide specificities of CRISPR-Cas9 nucleases[J]. Nature Reviews Genetics, 2016, 17(5):300-312.
[55] Lee CM, Cradick TJ, Fine EJ, et al.Nuclease target site selection for maximizing on-target activity and minimizing off-target effects in genome editing[J]. Molecular Therapy:the Journal of the American Society of Gene Therapy, 2016, 24(3):475-487.
[56] Osborn MJ, Webber BR, Knipping F, et al.Evaluation of TCR gene editing achieved by TALENs, CRISPR/Cas9, and megaTAL nucleases[J]. Molecular Therapy:the Journal of the American Society of Gene Therapy, 2016, 24(3):570-581.
[57] Tsai SQ, Zongni Z, Nguyen NT, et al.GUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases[J]. Nat Biotechnol, 2015, 33(2):187-197.
[58] Kim D, Kim S, Kim S, et al.Genome-wide target specificities of CRISPR-Cas9 nucleases revealed by multiplex Digenome-seq[J]. Genome Res, 2016, 26(3):406-415.
[59] Tsai SQ, Nguyen NT, Malagon Lopez J, et al.CIRCLE-seq:a highly sensitive in vitro screen for genome-wide CRISPR-Cas9 nuclease off-targets[J]. Nat Methods, 2017, 14(6):607-614.
[60] Cameron P, Fuller CK, Donohoue PD, et al.Mapping the genomic landscape of CRISPR-Cas9 cleavage[J]. Nat Methods, 2017, 14(6):600-606.
[61] Daesik K, Sangsu B, Jeongbin P, et al.Digenome-seq:genome-wide profiling of CRISPR-Cas9 off-target effects in human cells[J]. Nature Methods, 2015, 12(3):237-243.
[62] Wienert B, Wyman SK, Richardson CD, et al.Unbiased detection of CRISPR off-targets in vivo using DISCOVER-Seq[J]. Science, 2019, 364(6437):286-289.