Skip to main content

Genome Editing in Apple

  • Chapter
  • First Online:
The Apple Genome

Part of the book series: Compendium of Plant Genomes ((CPG))

Abstract

Genome editing with artificially engineered nucleases is an advanced molecular technology for pursuing precise and effective genetic engineering. In this technology, engineered nucleases induce DNA double-strand breaks at targeted sites in a genome, stimulating the DNA repair system in cells, thus enabling site-directed mutagenesis. Genome editing using CRISPR (clustered regularly interspaced short palindromic repeats)/CRISPR-associated protein9 (Cas9), originating from a defence system of prokaryotes, is a powerful technology that is now being widely utilized in molecular research studies, as well as in breeding programmes of various plant species, including fruit trees, to impart either novel or enhanced traits to established commercial cultivars or to new cultivars/genotypes. Recently, several reports have demonstrated successful apple genome editing and the introduction of important traits, such as those for early flowering and reduced fire blight susceptibility, to popular commercial cultivars, such as ‘Gala’ and ‘Golden Delicious’. It is important to point out that these reports reveal that such genome-edited/mutant apple plants or cell lines do not carry foreign genes. Nevertheless, during the process of precise genome editing, the coexistence of various types of mutations referred to as “mosaic mutations” and off-target effects are major concerns. Therefore, to minimize such effects, selection of target sequences and estimation of off-target effects for CRISPR/Cas9 has been developed for many organisms, and these have also been employed for apple by using in silico analysis based on genome information. On the other hand, apple genome heterozygosity has led to difficulties in genome editing, as the complex genome of apple precludes the use of some of these basic techniques for genome editing. Therefore, further studies focused on genome information and culture techniques tailored for apple are needed. It will be highly critical for each apple cultivar in developing precise and efficient genome editing for apple. This chapter will provide an overview of current studies of genome editing in apple and will discern and explore how this strategy will provide insights into molecular breeding technologies for genetic improvement of the apple.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 189.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 249.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  • Abu-Qaoud H, Skirvin RM, Chevreau E (1990) In vitro separation of chimeral pears into their component genotypes. Euphytica 48(2):189–196

    Article  Google Scholar 

  • Andersson M, Turesson H, Olsson N, Fält AS, Ohlsson P, Gonzalez MN, Samuelsson M, Hofvander P (2018) Genome editing in potato via CRISPR-Cas9 ribonucleoprotein delivery. Physiol Plant 164(4):378–384

    Article  CAS  PubMed  Google Scholar 

  • Borejsza-Wysocka EE, Malnoy M, Meng X, Bonasera JM, Nissinen RM, Kim JF, Beer SV, Aldwinckle HS (2004) Silencing of apple proteins that interact with DspE, a pathogenicity effector from Erwinia amylovora, as a strategy to increase resistance to fire blight Acta Hortic 663:469–474

    Google Scholar 

  • Bortesi L, Fischer R (2015) The CRISPR/Cas9 system for plant genome editing and beyond. Biotech Adv 33(1):41–52

    Article  CAS  Google Scholar 

  • Broothaerts W, Keulemans J, Van Nerum I (2004) Self-fertile apple resulting from S-RNase gene silencing. Plant Cell Rep 22(7):497–501

    Article  CAS  PubMed  Google Scholar 

  • Čermák T, Baltes NJ, Čegan R, Zhang Y, Voytas DF (2015) High-frequency, precise modification of the tomato genome. Genome Biol 16(1):232. https://doi.org/10.1186/s13059-015-0796-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Charrier A, Vergne E, Dousset N, Richer A, Petiteau A, Chevreau E (2019) Efficient targeted mutagenesis in apple and first time edition of pear using the CRISPR-Cas9 system. Front Plant Sci 10:40. https://doi.org/10.3389/fpls.2019.00040

  • Chen X, Li S, Zhang D, Han M, Jin X, Zhao C, Wang S, Xing L, Ma J, Ji J, An N (2019) Sequencing of a wild apple (Malus baccata) genome unravels the differences between cultivated and wild apple species regarding disease resistance and cold tolerance. G3: Genes Genomes Genet 9(7):2051–2060

    Google Scholar 

  • Clasen BM, Stoddard TJ, Luo S, Demorest ZL, Li J, Cedrone F, Tibebu R, Davison S, Ray EE, Daulhac A, Coffman A, Yabandith A, Retterath A, Haun W, Baltes NJ, Mathis L, Voytas DF, Zhang F (2016) Improving cold storage and processing traits in potato through targeted gene knockout. Plant Biotech J 14(1):169–176

    Article  CAS  Google Scholar 

  • Crosby JA, Janick J, Pecknold PC, Goffreda JC, Korban SS (1994) ‘GoldRush’ apple. HortScience 29(7):827–828

    Article  Google Scholar 

  • Daccord N, Celton JM, Linsmith G, Becker C, Choisne N, Schijlen E, van de Geest H, Bianco L, Micheletti D, Velasco R, Pierro EAD, Gouzy J, Rees DJG, Guérif P, Muranty H, Durel CE, Laurens F, Lespinasse Y, Gaillard S, Aubourg S, Quesneville H, Weigel D, van de Weg E, Troggio M, Bucher E (2017) High-quality de novo assembly of the apple genome and methylome dynamics of early fruit development. Nat Genet 49(7):1099–1106. https://doi.org/10.1038/ng.3886

    Article  CAS  PubMed  Google Scholar 

  • Dahan-Meir T, Filler-Hayut S, Melamed-Bessudo C, Bocobza S, Czosnek H, Aharoni A, Levy AA (2018) Efficient in planta gene targeting in tomato using geminiviral replicons and the CRISPR/Cas9 system. Plant J 95(1):5–16

    Article  CAS  PubMed  Google Scholar 

  • Fu Y, Sander JD, Reyon D, Cascio VM, Joung JK (2014) Improving CRISPR-Cas nuclease specificity using truncated guide RNAs. Nat Biotech 32(3):279–284

    Article  CAS  Google Scholar 

  • Gao C (2019) Precision plant breeding using genome editing technologies. Transgenic Res 28:53–55

    Article  CAS  PubMed  Google Scholar 

  • Gil-Humanes J, Wang Y, Liang Z, Shan Q, Ozuna CV, Sánchez-León S, Baltes NJ, Starker C, Barro F, Gao C, Voytas DF (2017) High-efficiency gene targeting in hexaploid wheat using DNA replicons and CRISPR/Cas9. Plant J 89(6):1251–1262

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Haeussler M, Schönig K, Eckert H, Eschstruth A, Mianné J, Renaud JB, Schneider-Maunoury S, Shkumatava A, Teboul L, Kent J, Joly JS, Concordet JP (2016) Evaluation of off-target and on-target scoring algorithms and integration into the guide RNA selection tool CRISPOR. Genome Biol 17(1):148. https://doi.org/10.1186/s13059-016-1012-2

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hayut SF, Bessudo CM, Levy AA (2017) Targeted recombination between homologous chromosomes for precise breeding in tomato. Nat Commun 8(1):15605. https://doi.org/10.1038/ncomms15605

    Article  CAS  Google Scholar 

  • Iwata H, Minamikawa MF, Kajiya-Kanegae H, Ishimori M, Hayashi T (2016) Genomics-assisted breeding in fruit trees. Breed Sci 66(1):100–115

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • James DJ, Passey AJ, Barbara DJ, Bevan M (1989) Genetic transformation of apple (Malus pumila Mill.) using a disarmed Ti-binary vector. Plant Cell Rep 7(8):658–661

    Google Scholar 

  • Kamburova VS, Nikitina EV, Shermatov SE, Buriev ZT, Kumpatla SP, Emani C, Abdurakhmonov IY (2017) Genome editing in plants: an overview of tools aapplications. Int J Agron 2017: Article ID 7315351. https://doi.org/10.1155/2017/7315351

  • Kaur N, Alok A, Shivani KN, Pandey P, Awasthi P, Tiwari S (2018) CRISPR/Cas9-mediated efficient editing in phytoene desaturase (PDS) demonstrates precise manipulation in banana cv. Rasthali Genome Funct Integr Genomics 18(1):89–99

    Article  CAS  PubMed  Google Scholar 

  • Kim H, Kim ST, Ryu J, Kang BC, Kim JS, Kim SG (2017) CRISPR/Cpf1-mediated DNA-free plant genome editing. Nat Commun 8(1):14406. https://doi.org/10.1038/ncomms14406

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kotoda N, Iwanami H, Takahashi S, Abe K (2006) Antisense expression of MdTFL1, a TFL1-like gene, reduces the juvenile phase in apple. J Am Soc Hortic Sci 131(1):74–81

    Article  CAS  Google Scholar 

  • Kunihisa M, Moriya S, Abe K, Okada K, Haji T, Hayashi T, Kawahara Y, Itoh R, Itoh T, Katayose Y, Kanamori H, Matsumoto T, Mori S, Sasaki H, Matsumoto T, Nishitani C, Terakami S, Yamamoto T (2016) Genomic dissection of a ‘Fuji’ apple cultivar: re-sequencing, SNP marker development, definition of haplotypes, and QTL detection. Breed Sci 66(4):499–515

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liang Z, Chen K, Li T, Zhang Y, Wang Y, Zhao Q, Liu J, Zhang H, Liu C, Ran Y, Gao C (2017) Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes. Nat Commun 8(1):14261. https://doi.org/10.1038/ncomms14261

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Limera C, Sabbadini S, Sweet JB, Mezzetti B (2017) New biotechnological tools for the genetic improvement of major woody fruit species. Front Plant Sci 8:1418. https://doi.org/10.3389/fpls.2017.01418

    Article  PubMed  PubMed Central  Google Scholar 

  • Liu H, Ding Y, Zhou Y, Jin W, Xie K, Chen LL (2017) CRISPR-P 2.0: an improved CRISPR-Cas9 tool for genome editing in plants. Mol Plant 10(3):530–532

    Google Scholar 

  • Liu X, Homma A, Sayadi J, Yang S, Ohashi J, Takumi T (2016) Sequence features associated with the cleavage efficiency of CRISPR/Cas9 system. Sci Rep 6:19675. https://doi.org/10.1038/srep19675

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Malnoy M, Viola R, Jung MH, Koo OJ, Kim S, Kim JS, Velasco R, Nagamangala Kanchiswamy C (2016) DNA-free genetically edited grapevine and apple protoplast using CRISPR/Cas9 ribonucleoproteins. Front Plant Sci 7:1904. https://doi.org/10.3389/fpls.2016.01904

    Article  PubMed  PubMed Central  Google Scholar 

  • Mehravar M, Shirazi A, Nazari M, Banan M (2019) Mosaicism in CRISPR/Cas9-mediated genome editing. Dev Biol 445(2):156–162

    Article  CAS  PubMed  Google Scholar 

  • Miki D, Zhang W, Zeng W, Feng Z, Zhu JK (2018) CRISPR/Cas9-mediated gene targeting in Arabidopsis using sequential transformation. Nat Commun 9(1):1967. https://doi.org/10.1038/s41467-018-04416-0

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Milčevičová R, Gosch C, Halbwirth H, Stich K, Hanke MV, Peil A, Flachowsky H, Rozhon W, Jonak C, Oufir M, Hausman JF, Matušíková I, Fluch S, Wilhelm E (2010) Erwinia amylovora-induced defense mechanisms of two apple species that differ in susceptibility to fire blight. Plant Sci 179(1–2):60–67

    Article  Google Scholar 

  • Mimida N, Kotoda N, Ueda T, Igarashi M, Hatsuyama Y, Iwanami H, Moriya S, Abe K (2009) Four TFL1/CEN-like genes on distinct linkage groups show different expression patterns to regulate vegetative and reproductive development in apple (Malus× domestica Borkh.). Plant Cell Physiol 50(2): 394–412

    Google Scholar 

  • Murata M, Nishimura M, Murai N, Haruta M, Homma S, Itoh Y (2001) A transgenic apple callus showing reduced polyphenol oxidase activity and lower browning potential. Biosci Biotech Biochem 65(2):383–388

    Article  CAS  Google Scholar 

  • Nakajima I, Ban Y, Azuma A, Onoue N, Moriguchi T, Yamamoto T, Toki S, Endo M (2017) CRISPR/Cas9-mediated targeted mutagenesis in grape. PLoS ONE 12(5):e0177966. https://doi.org/10.1371/journal.pone.0177966

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Nishitani C, Hirai N, Komori S, Wada M, Okada K, Osakabe K, Yamamoto T, Osakabe Y (2016) Efficient genome editing in apple using a CRISPR/Cas9 system. Sci Rep 6:31481. https://doi.org/10.1038/srep31481

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Norelli JL, Aldwinckle HS, Destéfano-Beltrán L, Jaynes JM (1994) Transgenic ‘Mailing 26’apple expressing the attacin E gene has increased resistance to Erwinia amylovora. Euphytica 77(1–2):123–128

    Article  CAS  Google Scholar 

  • Ohmori M, Yamane H, Osakabe K, Osakabe Y, Tao R (2020) Targeted mutagenesis of CENTRORADIALIS using CRISPR/Cas9 system through the improvement of genetic transformation efficiency of tetraploid highbush blueberry. J Hortic Sci Biotech. https://doi.org/10.1080/14620316.2020.1822760

    Article  Google Scholar 

  • Osakabe Y, Osakabe K (2015) Genome editing with engineered nucleases in plants. Plant Cell Physiol 56(3):389–400

    Article  CAS  PubMed  Google Scholar 

  • Osakabe Y, Watanabe T, Sugano SS, Ueta R, Ishihara R, Shinozaki K, Osakabe K (2016) Optimization of CRISPR/Cas9 genome editing to modify abiotic stress responses in plants. Sci Rep 6:26685. https://doi.org/10.1038/srep26685

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Osakabe Y, Liang Z, Ren C, Nishitani C, Osakabe K, Wada M, Komori S, Malnoy M, Velasco R, Poli M, Jung MH, Koo OJ, Viola R, Kanchiswamy CN (2018) CRISPR–Cas9-mediated genome editing in apple and grapevine. Nat Protoc 13(12):2844–2863

    Article  CAS  PubMed  Google Scholar 

  • Pan C, Ye L, Qin L, Liu X, He Y, Wang J, Chen L, Lu G (2016) CRISPR/Cas9-mediated efficient and heritable targeted mutagenesis in tomato plants in the first and later generations. Sci Rep 6:24765. https://doi.org/10.1038/srep24765

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Peer R, Rivlin G, Golobovitch S, Lapidot M, Gal-On A, Vainstein A, Tzfira T, Flaishman MA (2015) Targeted mutagenesis using zinc-finger nucleases in perennial fruit trees. Planta 241(4):941–951

    Article  CAS  PubMed  Google Scholar 

  • Peng A, Chen S, Lei T, Xu L, He Y, Wu L, Yao L, Zou X (2017) Engineering canker-resistant plants through CRISPR/Cas9-targeted editing of the susceptibility gene CsLOB1 promoter in citrus. Plant Biotech J 15(12):1509–1519

    Article  CAS  Google Scholar 

  • Pompili V, Dalla Costa L, Piazza S, Pindo M, Malnoy M (2020) Reduced fire blight susceptibility in apple cultivars using a high-efficiency CRISPR/Cas9-FLP/FRT-based gene editing system. Plant Biotech J 18(3):845–858. https://doi.org/10.1111/pbi.13253

    Article  CAS  Google Scholar 

  • Puchta H, Fauser F (2013) Gene targeting in plants: 25 years later. Int J Dev Biol 57:629–637

    Article  CAS  PubMed  Google Scholar 

  • Puite KJ, Schaart JG (1996) Genetic modification of the commercial apple cultivars Gala, Golden Delicious and Elstar via an Agrobacterium tumefaciens-mediated transformation method. Plant Sci 119:125–133

    Article  CAS  Google Scholar 

  • Ren C, Liu X, Zhang Z, Wang Y, Duan W, Li S, Liang Z (2016) CRISPR/Cas9-mediated efficient targeted mutagenesis in Chardonnay (Vitis vinifera L.). Sci Rep 6:32289. https://doi.org/10.1038/srep32289

  • Rong YS, Golic KG (2003) The homologous chromosome is an effective template for the repair of mitotic DNA double-strand breaks in Drosophila. Genetics 165(4):1831–1842

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rozov SM, Permyakova NV, Deineko EV (2019) The problem of the low rates of CRISPR/Cas9-mediated knock-ins in plants: Approaches and solutions. Int J Mol Sci 20(13):3371. https://doi.org/10.3390/ijms20133371

    Article  CAS  PubMed Central  Google Scholar 

  • Seong ES, Song KJ (2008) Factors affecting the early gene transfer step in the development of transgenic ‘Fuji’ apple plants. Plant Grow Reg 54(2):89–95

    Article  CAS  Google Scholar 

  • Shannon S, Meeks-Wagner DR (1991) A mutation in the Arabidopsis TFL1 gene affects inflorescence meristem development. Plant Cell 3(9):877–892

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Spiegel-Roy P (1990) Economic and agricultural impact of mutation breeding in fruit trees. Mut Breed Rev 5:1–26

    Google Scholar 

  • Steinert J, Schiml S, Puchta H (2016) Homology-based double-strand break-induced genome engineering in plants. Plant Cell Rep 35(7):1429–1438

    Article  CAS  PubMed  Google Scholar 

  • Subburaj S, Chung SJ, Lee C, Ryu SM, Kim DH, Kim JS, Bae S, Lee GJ (2016) Site-directed mutagenesis in Petunia × hybrida protoplast system using direct delivery of purified recombinant Cas9 ribonucleoproteins. Plant Cell Rep 35(7):1535–1544

    Article  CAS  PubMed  Google Scholar 

  • Svitashev S, Schwartz C, Lenderts B, Young JK, Cigan AM (2016) Genome editing in maize directed by CRISPR–Cas9 ribonucleoprotein complexes. Nat Commun 7(1):13274. https://doi.org/10.1038/ncomms13274

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ueta R, Abe C, Watanabe T, Sugano SS, Ishihara R, Ezura H, Osakabe Y, Osakabe K (2017) Rapid breeding of parthenocarpic tomato plants using CRISPR/Cas9. Sci Rep 7:507. https://doi.org/10.1038/s41598-017-00501-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Velasco R, Zharkikh A, Affourtit J, Dhingra A, Cestaro A, Kalyanaraman A, Fontana P, Bhatnagar SK, Troggio M, Pruss D, Salvi S, Pindo M, Baldi P, Castelletti S, Cavaiuolo M, Coppola G, Costa F, Cova V, Ri AD, Goremykin V, Komjanc M, Longhi S, Magnago P, Malacarne G, Malnoy M, Micheletti D, Moretto M, Perazzolli M, Si-Ammour A, Vezzulli S, Zini E, Eldredge G, Fitzgerald LM, Gutin N, Lanchbury G, Macalma T, Mitchell JT, Reid J, Wardell B, Kodira C, Chen Z, Desany B, Niazi F, Palmer M, Koepke T, Jiwan D, Schaeffer S, Krishnan V, Wu C, Chu VT, King ST, Vick J, Tao Q, Mraz A, Stormo A, Stormo K, Bogden R, Ederle D, Stella A, Vecchietti A, Kater MM, Masiero S, Lasserre P, Lespinasse Y, Allan AC, Bus V, Chagné D, Crowhurst RN, Gleave AP, Lavezzo E, Fawcett JA, Proost S, Rouzé P, Sterck L, Toppo S, Lazzari B, Hellens RP, Durel C-E, Gutin A, Bumgarner R, Gardiner SE, Skolnick M, Egholm M, Van de Peer Y, Salamini F, Viola R (2010) The genome of the domesticated apple (Malus × domestica Borkh.). Nat Genet 42:833–839

    Article  CAS  PubMed  Google Scholar 

  • Wada M, Ureshino A, Tanaka N, Komori S, Takahashi S, Kudo K, Bessho H (2009) Anatomical analysis by two approaches ensure the promoter activities of apple AFL genes. J Jpn Soc Hortic Sc 78(1):32–39

    Article  CAS  Google Scholar 

  • Wang M, Lu Y, Botella JR, Mao Y, Hua K, Zhu JK (2017) Gene targeting by homology-directed repair in rice using a geminivirus-based CRISP)R/Cas9 system. Mol Plant 10(7):1007–1010

    Article  CAS  PubMed  Google Scholar 

  • Wang Z, Wang S, Li D, Zhang Q, Li L, Zhong C, Liu Y, Huang H (2018) Optimized paired-sgRNA/Cas9 cloning and expression cassette triggers high-efficiency multiplex genome editing in kiwifruit. Plant Biotech J 16(8):1424–1433

    Article  CAS  Google Scholar 

  • Woo JW, Kim J, Kwon SI, Corvalán C, Cho SW, Kim H, Kim SG, Kim ST, Choe S, Kim JS (2015) DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleoproteins. Nat Biotech 33(11):1162–1164

    Article  CAS  Google Scholar 

  • Xiao A, Cheng Z, Kong L, Zhu Z, Lin S, Gao G, Zhang B (2014) CasOT: a genome-wide Cas9/gRNA off-target searching tool. Bioinformatics 30(8):1180–1182

    Article  CAS  PubMed  Google Scholar 

  • Zhang L, Hu J, Han X, Li J, Gao Y, Richards CM, Zhang C, Tian Y, Liu G, Gul H, Wang D, Tian Y, Yang C, Meng M, Yuan G, Kang G, Wu Y, Wang K, Zhang H, Wang D, Cong P (2019) A high-quality apple genome assembly reveals the association of a retrotransposon and red fruit colour. Nat Commun 10:1494. https://doi.org/10.1038/s41467-019-09518-x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Yuriko Osakabe .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2021 Springer Nature Switzerland AG

About this chapter

Check for updates. Verify currency and authenticity via CrossMark

Cite this chapter

Nishitani, C., Osakabe, K., Osakabe, Y. (2021). Genome Editing in Apple. In: Korban, S.S. (eds) The Apple Genome. Compendium of Plant Genomes. Springer, Cham. https://doi.org/10.1007/978-3-030-74682-7_10

Download citation

Publish with us

Policies and ethics