CRISPR/Cas9基因编辑技术在果蔬作物中的应用
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摘要
成簇的规律间隔的短回文重复序列(clustered regularly interspaced short palindromic repeats, CRISPR)及其相关蛋白(CRISPR-associated, Cas)是存在于细菌和古细菌中的获得性免疫系统,可以对外来某一特定单链或双链DNA实施高特异性地沉默或降解。近年来, CRISPR/Cas9系统已快速发展为一种基因组定向编辑技术。由于该技术操作简单、快速、突变诱导率高等优点,已在多种果蔬作物中实现了基因的编辑,在果蔬作物性状改良、基因表达、代谢过程调控及逆境胁迫抗性相关基因的挖掘等研究中发挥了重要作用。本文就CRISPR/Cas9系统的结构、作用原理、在果蔬作物基因编辑研究中的应用及存在的问题等进行了分析和总结。
Clustered regularly interspaced short palindromic repeats(CRISPR) and CRISPR-associated protein(Cas) is a prokaryotic immune system which derived from bacteria and archaea, and can be used to silence or degrade a specific foreign single-stranded or double-stranded DNA with high specificity. In recent years, the CRISPR/Cas9 system has been rapidly developed as a genome-oriented editing technology. Due to its simple, rapid, and high mutation induction rate, the technology has achieved gene editing in a variety of fruit and vegetable crops, and plays important roles in the improvement of fruit and vegetable characters, gene expression, metabolic process regulation and stress resistance related gene mining. In this review, we introduced the structure and principle of CRISPR/Cas9 system and summarized the applications of this system in fruits and vegetables. Meanwhile, the problems and its solutions of the application of CRISPR/Cas9 system were also analyzed.
Clustered regularly interspaced short palindromic repeats(CRISPR) and CRISPR-associated protein(Cas) is a prokaryotic immune system which derived from bacteria and archaea, and can be used to silence or degrade a specific foreign single-stranded or double-stranded DNA with high specificity. In recent years, the CRISPR/Cas9 system has been rapidly developed as a genome-oriented editing technology. Due to its simple, rapid, and high mutation induction rate, the technology has achieved gene editing in a variety of fruit and vegetable crops, and plays important roles in the improvement of fruit and vegetable characters, gene expression, metabolic process regulation and stress resistance related gene mining. In this review, we introduced the structure and principle of CRISPR/Cas9 system and summarized the applications of this system in fruits and vegetables. Meanwhile, the problems and its solutions of the application of CRISPR/Cas9 system were also analyzed.
引文
Bao H,Chen X,LüS,et al(2015).Virus-induced gene silencing reveals control of reactive oxygen species accumulation and salt tolerance in tomato byγ-aminobutyric acid metabolic pathway.Plant Cell Environ,38(3):600-613
Bedell VM,Wang Y,Campbell JM,et al(2012).In vivo genome editing using high efficiency TALENs.Nature,491:114-118
Bhaya D,Davison M,Barrangou R(2011).CRISPR-Cas systems in bacteria and archaea:versatile small RNAs for adaptive defense and regulation.Annu Rev Genet,45(45):273-297
Brooks C,Nekrasov V,Lippman ZB,et al(2014).Efficient gene editing in tomato in the first generation using the clustered regularly interspaced short palindromic repeats/CRISPR-associated9 system.Plant Physiol,166(3):1292-1297
Butler NM,Atkins PA,Voytas DF,et al(2015).Generation and inheritance of targeted mutations in potato(Solanum tuberosum L.)using the CRISPR/Cas system.PLoS One,10:e0144591
Cao HL,Chen D,Ye NX,et al(2017).Cloning and abiotic stress expression analysis of CsMAPK3 gene in tea plant.Acta Hortic Sin,44(11):2203-2214(in Chinese with English abstract)[曹红利,陈丹,叶乃兴等(2017).茶树CsMAPK3的全长克隆及其逆境表达分析.园艺学报,44(11):2203-2214]
Carroll D(2011).Genome engineering with zinc-finger nucleases.Genetics,188:773-782
Chandrasekaran J,Brumin M,Wolf D,et al(2016).Development of broad virus resistance in non-transgenic cucumber using CRISPR/Cas9 technology.Mol Plant Pathol,17(7):1140-1153
Chen JS,Dagdas YS,Kleinstiver BP,et al(2017).Enhanced proofreading governs CRISPR-Cas9 targeting accuracy.Nature,550(7676):407-410
Christian M,Cermak T,Doyle EL,et al(2010).Targeting DNA double-strand breaks with TAL effector nucleases.Genetics,186:757-761
Cradick TJ,Qiu P,Lee CM,et al(2014).COSMID:A webbased tool for identifying and validating CRISPR/Cas off-target sites.Mol Ther-Nucl Acids,3(12):e214
D’Ambrosio C,Stigliani AL,Giorio G(2018).CRISPR/Cas9editing of carotenoid genes in tomato.Transgenic Res,27(4):367-378
Deltcheva E,Chylinski K,Sharma CM,et al(2011).CRISPRRNA maturation by trans-encoded small RNA and host factor RNase III.Nature,471(7340):602-607
Deng L,Wang H,Sun C,et al(2018).Efficient generation of pink-fruited tomatoes using CRISPR/Cas9 system.J Genet Genomics,45(1):51-54
Doudna JA,Charpentier E(2014).The new frontier of genome engineering with CRISPR-Cas9.Science,346(6213):1258096
Grissa I,Vergnaud G,Pourcel C(2007).The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats.BMC Bioinformatics,8(1):172-182
Guilinger JP,Thompson DB,Liu DR(2014).Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification.Nat Biotechnol,32(6):577-582
Hsu PD,Scott DA,Weinstein JA,et al(2013).DNA targeting specificity of RNA-guided Cas9 nucleases.Nat Biotechnol,31(9):827-832
Hu JH,Miller SM,Geurts MH,et al(2018).Evolved Cas9variants with broad PAM compatibility and high DNAspecificity.Nature,556(7699):57-63
Hua K,Tao X,Zhu JK(2019).Expanding the base editing scope in rice by using Cas9 variants.Plant Biotechnol J,17(2):499-504
Huang X,Tang L,Yu Y,et al(2018).Control of flowering and inflorescence architecture in tomato by synergistic interactions between ALOG transcription factors.J Genet Genomics,45(10):557-560
Jia H,Zhang Y,Orbovi?V,et al(2017).Genome editing of the disease susceptibility gene CsLOB1 in citrus confers resistance to citrus canker.Plant Biotechnol J,15(7):817-823
Kaur N,Alok A,Shivani,et al(2018).CRISPR/Cas9-mediated efficient editing in phytoene desaturase(PDS)demonstrates precise manipulation in banana cv.Rasthali genome.Funct Integr Genomic,18(1):89-99
Kleinstiver BP,Pattanayak V,Prew MS,et al(2016).High-fidelity CRISPR-Cas9 nucleases with no detectable genome-wide off-target effects.Nature,529(7587):490-495
Kleinstiver BP,Prew MS,Tsai SQ,et al(2015).Engineered CRISPR-Cas9 nucleases with altered PAM specificities.Nature,523(7561):481-485
Klimek-Chodacka M,Oleszkiewicz T,Lowder LG,et al(2018).Efficient CRISPR/Cas9-based genome editing in carrot cells.Plant Cell Rep,37(4):575-586
Koike S,Matsukura C,Takayama M,et al(2013).Suppression ofγ-aminobutyric acid(GABA)transaminases induces prominent GABA accumulation,dwarfism and infertility in the tomato(Solanum lycopersicum L.).Plant Cell Physiol,54(5):793-807
Lawrenson T,Shorinola O,Stacey N,et al(2015).Induction of targeted,heritable mutations in barley and brassica oleracea using RNA-guided Cas9 nuclease.Genome Biol,16(1):258-271
Li JF,Aach J,Norville JE,et al(2013).Multiplex and homologous recombination-mediated genome editing in Arabidopsis and Nicotiana benthamiana using guide RNA and Cas9.Nat Biotechnol,31(8):688-691
Li R,Fu D,Zhu B,et al(2018a).CRISPR/Cas9-mediated mutagenesis of lncRNA1459 alters tomato fruit ripening.Plant J,94(3):513-524
Li R,Li R,Li X,et al(2017).Multiplexed CRISPR/Cas9-mediated metabolic engineering ofγ-aminobutyric acid levels in Solanum lycopersicum.Plant Biotechnol J,16(2):415-427
Li S,Zhang X,Wang W,et al(2018c).Expanding the scope of CRISPR/Cpf1-mediated genome editing in rice.Mol Plant,11(7):995-998
Li X,Wang Y,Chen S,et al(2018b).Lycopene is enriched in tomato fruit by CRISPR/Cas9-mediated multiplex genome editing.Front Plant Sci,9:559
Liang Z,Chen K,Li T,et al(2017).Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes.Nat Commun,8:14261
Lu Y,Zhu JK(2017).Precise editing of a target base in the rice genome using a modified CRISPR/Cas9 System.Mol Plant,10(3):523-525
Ma X,Zhu Q,Chen Y,et al(2016).CRISPR/Cas9 platforms for genome editing in plants:developments and applications.Mol Plant,9(7):961-974
Ma XL,Liu YG(2016).CRISPR/Cas9-based genome editing systems and the analysis of targeted genome mutations in plants.Hereditas,38(2):118-125(in Chinese with English abstract)[马兴亮,刘耀光(2016).植物CRISPR/Cas9基因组编辑系统与突变分析.遗传,38(2):118-125]
Mahfouz MM,Tashkandi M,Ali Z,et al(2018).Engineering resistance against Tomato yellow leaf curl virus via the CRISPR/Cas9 system in tomato.Plant Signal Behav,13(10):e1525996
Makarova KS,Zhang F,Koonin EV,et al(2017a).SnapShot:class 2 CRISPR-Cas systems.Cell,168(1):328-328.e1
Makarova KS,Zhang F,Koonin EV,et al(2017b).SnapShot:class 1 CRISPR-Cas systems.Cell,168(5):946-946.e1
Malnoy M,Viola R,Jung MH,et al(2016).DNA-free genetically edited grapevine and apple protoplast using CRIS-PR/Cas9 ribonucleoproteins.Front Plant Sci,7:1904
Mikami M,Toki S,Endo M,et al(2015).Comparison of CRISPR/Cas9 expression constructs for efficient targeted mutagenesis in rice.Plant Mol Biol,88(6):561-572
Nekrasov V,Staskawicz B,Weigel D,et al(2013).Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease.Nat Biotechnol,31(8):691-693
Nekrasov V,Wang C,Win J,et al(2017).Rapid generation of a transgene-free powdery mildew resistant tomato by genome deletion.Sci Rep,7(1):482-488
Nishitani C,Hirai N,Komori S,et al(2016).Efficient genome editing in apple using a CRISPR/Cas9 system.Sci Rep,6:31481
Nonaka S,Arai C,Takayama M,et al(2017).Efficient increase ofγ-aminobutyric acid(GABA)content in tomato fruits by targeted mutagenesis.Sci Rep,7(1):7057-7071
Parkhi V,Bhattacharya A,Choudhary S,et al(2018).Demonstration of CRISPR-cas9-mediated pds gene editing in a tomato hybrid parental line.Indian J Genet,78(1):132-137
Pattanayak V,Lin S,Guilinger JP,et al(2013).High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity.Nat Biotechnol,31(9):839-843
Peng A,Chen S,Lei T,et al(2017).Engineering canker-resistant plants through CRISPR/Cas9-targeted editing of the susceptibility gene CsLOB1 promoter in citrus.Plant Biotechnol J,15(12):1509-1519
Pougach K,Semenova EE,Datsenko KA,et al(2010).Transcription,processing and function of CRISPR cassettes in Escherichia coli.Mol Microbiol,77(6):1367-1379
Ran FA,Hsu PD,Lin CY,et al(2013).Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity.Cell,154(6):1380-1389
Ren C,Liu X,Zhang Z,et al(2016).CRISPR/Cas9-mediated efficient targeted mutagenesis in Chardonnay(Vitis vinifera L.).Sci Rep,6:32289
Sander JD,Joung JK(2014).CRISPR-Cas systems for editing,regulating and targeting genomes.Nat Biotechnol,32(4):347-355
Shan Q,Wang Y,Li J,et al(2013).Targeted genome modification of crop plants using a CRISPR-Cas system.Nat Biotechnol,31(8):686-688
Snowden CJ,Thomas B,Baxter CJ,et al(2015).A tonoplast Glu/Asp/GABA exchanger that affects tomato fruit amino acid composition.Plant J,81(5):651-660
Soyk S,Müller NA,Park SJ,et al(2017).Variation in the flowering gene SELF PRUNING 5G promotes day-neutrality and early yield in tomato.Nat Genet,49(1):162-168
Sternberg SH,Redding S,Jinek M,et al(2014).DNA interrogation by the CRISPR RNA-guided endonuclease Cas9.Nature,507(7490):62-67
Takayama M,Ezura H(2015).How and why does tomato accumulate a large amount of GABA in the fruit?Front Plant Sci,6:612
Thomazella DP,Brail Q,Dahlbeck D,et al(2016).CRIS-PR-Cas9 mediated mutagenesis of a DMR6 ortholog in tomato confers broad-spectrum disease resistance.bioRxiv,doi:http://doi.org/10.1101/064824
Tian S,Jiang L,Cui X,et al(2018).Engineering herbicide-resistant watermelon variety through CRISPR/Cas9-mediated base-editing.Plant Cell Rep,37(9):1353-1356
Tian S,Jiang L,Gao Q,et al(2017).Efficient CRISPR/Cas9-based gene knockout in watermelon.Plant Cell Rep,36(3):399-406
Wang L,Chen L,Li R,et al(2017).Reduced drought tolerance by CRISPR/Cas9-mediated SlMAPK3 mutagenesis in tomato plants.J Agric Food Chem,65(39):8674-8682
Wang T,Birsoy K,Hughes NW,et al(2015).Identification and characterization of essential genes in the human genome.Science,350(6264):1096-1101
Watanabe K,Oda-Yamamizo C,Sage-Ono K,et al(2018).Alteration of flower colour in ipomoea nil through CRISPR/Cas9-mediated mutagenesis of carotenoid cleavage dioxygenase 4.Transgenic Res,27(1):25-38
Woo JW,Kim J,Kwon SI,et al(2015).DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleoproteins.Nat Biotechnol,33(11):1162-1164
Xie K,Zhang J,Yang Y(2014).Genome-wide prediction of highly specific guide RNA spacers for CRISPR-Cas9-mediated genome editing in model plants and major crops.Mol Plant,7(5):923-926
Xu C,Park SJ,Eck JV,et al(2016).Control of inflorescence architecture in tomato by BTB/POZ transcriptional regulators.Genes Dev,30(18):2048-2061
Yan M,Zhou SR,Xue HW(2015).CRISPR Primer Designer:Design primers for knockout and chromosome imaging CRISPR-Cas system.J Integr Plant Biol,57(7):613-617
Yifhar T,Pekker I,Peled D,et al(2012).Failure of the tomato trans-acting short interfering RNA program to regulate auxin response factor3 and ARF4 underlies the wiry leaf syndrome.Plant Cell,24(9):3575-3589
Zeilmaker T,Ludwig NR,Elberse J,et al(2015).Downy mildew resistant 6 and DMR6-LIKE oxygenase 1 are partially redundant but distinct suppressors of immunity in Arabidopsis.Plant J,81(2):210-222
Zetsche B,Gootenberg JS,Abudayyeh OO,et al(2015).Cpf1is a single RNA-guided endonuclease of a class 2 CRIS-PR-Cas system.Cell,163(3):759-771
Zhang B,Yang X,Yang C,et al(2016).Exploiting the CRIS-PR/Cas9 system for targeted genome mutagenesis in petunia.Sci Rep,6:20315
Bedell VM,Wang Y,Campbell JM,et al(2012).In vivo genome editing using high efficiency TALENs.Nature,491:114-118
Bhaya D,Davison M,Barrangou R(2011).CRISPR-Cas systems in bacteria and archaea:versatile small RNAs for adaptive defense and regulation.Annu Rev Genet,45(45):273-297
Brooks C,Nekrasov V,Lippman ZB,et al(2014).Efficient gene editing in tomato in the first generation using the clustered regularly interspaced short palindromic repeats/CRISPR-associated9 system.Plant Physiol,166(3):1292-1297
Butler NM,Atkins PA,Voytas DF,et al(2015).Generation and inheritance of targeted mutations in potato(Solanum tuberosum L.)using the CRISPR/Cas system.PLoS One,10:e0144591
Cao HL,Chen D,Ye NX,et al(2017).Cloning and abiotic stress expression analysis of CsMAPK3 gene in tea plant.Acta Hortic Sin,44(11):2203-2214(in Chinese with English abstract)[曹红利,陈丹,叶乃兴等(2017).茶树CsMAPK3的全长克隆及其逆境表达分析.园艺学报,44(11):2203-2214]
Carroll D(2011).Genome engineering with zinc-finger nucleases.Genetics,188:773-782
Chandrasekaran J,Brumin M,Wolf D,et al(2016).Development of broad virus resistance in non-transgenic cucumber using CRISPR/Cas9 technology.Mol Plant Pathol,17(7):1140-1153
Chen JS,Dagdas YS,Kleinstiver BP,et al(2017).Enhanced proofreading governs CRISPR-Cas9 targeting accuracy.Nature,550(7676):407-410
Christian M,Cermak T,Doyle EL,et al(2010).Targeting DNA double-strand breaks with TAL effector nucleases.Genetics,186:757-761
Cradick TJ,Qiu P,Lee CM,et al(2014).COSMID:A webbased tool for identifying and validating CRISPR/Cas off-target sites.Mol Ther-Nucl Acids,3(12):e214
D’Ambrosio C,Stigliani AL,Giorio G(2018).CRISPR/Cas9editing of carotenoid genes in tomato.Transgenic Res,27(4):367-378
Deltcheva E,Chylinski K,Sharma CM,et al(2011).CRISPRRNA maturation by trans-encoded small RNA and host factor RNase III.Nature,471(7340):602-607
Deng L,Wang H,Sun C,et al(2018).Efficient generation of pink-fruited tomatoes using CRISPR/Cas9 system.J Genet Genomics,45(1):51-54
Doudna JA,Charpentier E(2014).The new frontier of genome engineering with CRISPR-Cas9.Science,346(6213):1258096
Grissa I,Vergnaud G,Pourcel C(2007).The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats.BMC Bioinformatics,8(1):172-182
Guilinger JP,Thompson DB,Liu DR(2014).Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification.Nat Biotechnol,32(6):577-582
Hsu PD,Scott DA,Weinstein JA,et al(2013).DNA targeting specificity of RNA-guided Cas9 nucleases.Nat Biotechnol,31(9):827-832
Hu JH,Miller SM,Geurts MH,et al(2018).Evolved Cas9variants with broad PAM compatibility and high DNAspecificity.Nature,556(7699):57-63
Hua K,Tao X,Zhu JK(2019).Expanding the base editing scope in rice by using Cas9 variants.Plant Biotechnol J,17(2):499-504
Huang X,Tang L,Yu Y,et al(2018).Control of flowering and inflorescence architecture in tomato by synergistic interactions between ALOG transcription factors.J Genet Genomics,45(10):557-560
Jia H,Zhang Y,Orbovi?V,et al(2017).Genome editing of the disease susceptibility gene CsLOB1 in citrus confers resistance to citrus canker.Plant Biotechnol J,15(7):817-823
Kaur N,Alok A,Shivani,et al(2018).CRISPR/Cas9-mediated efficient editing in phytoene desaturase(PDS)demonstrates precise manipulation in banana cv.Rasthali genome.Funct Integr Genomic,18(1):89-99
Kleinstiver BP,Pattanayak V,Prew MS,et al(2016).High-fidelity CRISPR-Cas9 nucleases with no detectable genome-wide off-target effects.Nature,529(7587):490-495
Kleinstiver BP,Prew MS,Tsai SQ,et al(2015).Engineered CRISPR-Cas9 nucleases with altered PAM specificities.Nature,523(7561):481-485
Klimek-Chodacka M,Oleszkiewicz T,Lowder LG,et al(2018).Efficient CRISPR/Cas9-based genome editing in carrot cells.Plant Cell Rep,37(4):575-586
Koike S,Matsukura C,Takayama M,et al(2013).Suppression ofγ-aminobutyric acid(GABA)transaminases induces prominent GABA accumulation,dwarfism and infertility in the tomato(Solanum lycopersicum L.).Plant Cell Physiol,54(5):793-807
Lawrenson T,Shorinola O,Stacey N,et al(2015).Induction of targeted,heritable mutations in barley and brassica oleracea using RNA-guided Cas9 nuclease.Genome Biol,16(1):258-271
Li JF,Aach J,Norville JE,et al(2013).Multiplex and homologous recombination-mediated genome editing in Arabidopsis and Nicotiana benthamiana using guide RNA and Cas9.Nat Biotechnol,31(8):688-691
Li R,Fu D,Zhu B,et al(2018a).CRISPR/Cas9-mediated mutagenesis of lncRNA1459 alters tomato fruit ripening.Plant J,94(3):513-524
Li R,Li R,Li X,et al(2017).Multiplexed CRISPR/Cas9-mediated metabolic engineering ofγ-aminobutyric acid levels in Solanum lycopersicum.Plant Biotechnol J,16(2):415-427
Li S,Zhang X,Wang W,et al(2018c).Expanding the scope of CRISPR/Cpf1-mediated genome editing in rice.Mol Plant,11(7):995-998
Li X,Wang Y,Chen S,et al(2018b).Lycopene is enriched in tomato fruit by CRISPR/Cas9-mediated multiplex genome editing.Front Plant Sci,9:559
Liang Z,Chen K,Li T,et al(2017).Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes.Nat Commun,8:14261
Lu Y,Zhu JK(2017).Precise editing of a target base in the rice genome using a modified CRISPR/Cas9 System.Mol Plant,10(3):523-525
Ma X,Zhu Q,Chen Y,et al(2016).CRISPR/Cas9 platforms for genome editing in plants:developments and applications.Mol Plant,9(7):961-974
Ma XL,Liu YG(2016).CRISPR/Cas9-based genome editing systems and the analysis of targeted genome mutations in plants.Hereditas,38(2):118-125(in Chinese with English abstract)[马兴亮,刘耀光(2016).植物CRISPR/Cas9基因组编辑系统与突变分析.遗传,38(2):118-125]
Mahfouz MM,Tashkandi M,Ali Z,et al(2018).Engineering resistance against Tomato yellow leaf curl virus via the CRISPR/Cas9 system in tomato.Plant Signal Behav,13(10):e1525996
Makarova KS,Zhang F,Koonin EV,et al(2017a).SnapShot:class 2 CRISPR-Cas systems.Cell,168(1):328-328.e1
Makarova KS,Zhang F,Koonin EV,et al(2017b).SnapShot:class 1 CRISPR-Cas systems.Cell,168(5):946-946.e1
Malnoy M,Viola R,Jung MH,et al(2016).DNA-free genetically edited grapevine and apple protoplast using CRIS-PR/Cas9 ribonucleoproteins.Front Plant Sci,7:1904
Mikami M,Toki S,Endo M,et al(2015).Comparison of CRISPR/Cas9 expression constructs for efficient targeted mutagenesis in rice.Plant Mol Biol,88(6):561-572
Nekrasov V,Staskawicz B,Weigel D,et al(2013).Targeted mutagenesis in the model plant Nicotiana benthamiana using Cas9 RNA-guided endonuclease.Nat Biotechnol,31(8):691-693
Nekrasov V,Wang C,Win J,et al(2017).Rapid generation of a transgene-free powdery mildew resistant tomato by genome deletion.Sci Rep,7(1):482-488
Nishitani C,Hirai N,Komori S,et al(2016).Efficient genome editing in apple using a CRISPR/Cas9 system.Sci Rep,6:31481
Nonaka S,Arai C,Takayama M,et al(2017).Efficient increase ofγ-aminobutyric acid(GABA)content in tomato fruits by targeted mutagenesis.Sci Rep,7(1):7057-7071
Parkhi V,Bhattacharya A,Choudhary S,et al(2018).Demonstration of CRISPR-cas9-mediated pds gene editing in a tomato hybrid parental line.Indian J Genet,78(1):132-137
Pattanayak V,Lin S,Guilinger JP,et al(2013).High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity.Nat Biotechnol,31(9):839-843
Peng A,Chen S,Lei T,et al(2017).Engineering canker-resistant plants through CRISPR/Cas9-targeted editing of the susceptibility gene CsLOB1 promoter in citrus.Plant Biotechnol J,15(12):1509-1519
Pougach K,Semenova EE,Datsenko KA,et al(2010).Transcription,processing and function of CRISPR cassettes in Escherichia coli.Mol Microbiol,77(6):1367-1379
Ran FA,Hsu PD,Lin CY,et al(2013).Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity.Cell,154(6):1380-1389
Ren C,Liu X,Zhang Z,et al(2016).CRISPR/Cas9-mediated efficient targeted mutagenesis in Chardonnay(Vitis vinifera L.).Sci Rep,6:32289
Sander JD,Joung JK(2014).CRISPR-Cas systems for editing,regulating and targeting genomes.Nat Biotechnol,32(4):347-355
Shan Q,Wang Y,Li J,et al(2013).Targeted genome modification of crop plants using a CRISPR-Cas system.Nat Biotechnol,31(8):686-688
Snowden CJ,Thomas B,Baxter CJ,et al(2015).A tonoplast Glu/Asp/GABA exchanger that affects tomato fruit amino acid composition.Plant J,81(5):651-660
Soyk S,Müller NA,Park SJ,et al(2017).Variation in the flowering gene SELF PRUNING 5G promotes day-neutrality and early yield in tomato.Nat Genet,49(1):162-168
Sternberg SH,Redding S,Jinek M,et al(2014).DNA interrogation by the CRISPR RNA-guided endonuclease Cas9.Nature,507(7490):62-67
Takayama M,Ezura H(2015).How and why does tomato accumulate a large amount of GABA in the fruit?Front Plant Sci,6:612
Thomazella DP,Brail Q,Dahlbeck D,et al(2016).CRIS-PR-Cas9 mediated mutagenesis of a DMR6 ortholog in tomato confers broad-spectrum disease resistance.bioRxiv,doi:http://doi.org/10.1101/064824
Tian S,Jiang L,Cui X,et al(2018).Engineering herbicide-resistant watermelon variety through CRISPR/Cas9-mediated base-editing.Plant Cell Rep,37(9):1353-1356
Tian S,Jiang L,Gao Q,et al(2017).Efficient CRISPR/Cas9-based gene knockout in watermelon.Plant Cell Rep,36(3):399-406
Wang L,Chen L,Li R,et al(2017).Reduced drought tolerance by CRISPR/Cas9-mediated SlMAPK3 mutagenesis in tomato plants.J Agric Food Chem,65(39):8674-8682
Wang T,Birsoy K,Hughes NW,et al(2015).Identification and characterization of essential genes in the human genome.Science,350(6264):1096-1101
Watanabe K,Oda-Yamamizo C,Sage-Ono K,et al(2018).Alteration of flower colour in ipomoea nil through CRISPR/Cas9-mediated mutagenesis of carotenoid cleavage dioxygenase 4.Transgenic Res,27(1):25-38
Woo JW,Kim J,Kwon SI,et al(2015).DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleoproteins.Nat Biotechnol,33(11):1162-1164
Xie K,Zhang J,Yang Y(2014).Genome-wide prediction of highly specific guide RNA spacers for CRISPR-Cas9-mediated genome editing in model plants and major crops.Mol Plant,7(5):923-926
Xu C,Park SJ,Eck JV,et al(2016).Control of inflorescence architecture in tomato by BTB/POZ transcriptional regulators.Genes Dev,30(18):2048-2061
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