Introduction
Fig. 1 Timeline of CRISPR/Cas9 system was first applied in different plant species |
Overview of CRISPR/Cas genome editing
Fig. 2 Schematics of genome editing technology systems and genome modifications generated by different systems. A ZFNs technology contains an array of engineered zinc finger proteins fused to catalytic domain of the FokI endonuclease. B TALENs have arrays of the TAL effector fused with FokI. C CRISPR/Cas9 system which is composed of Cas9 protein and sgRNA. (D) The cytidine base editor (CBE). Green circle represents cytidine deaminase rAPOBEC1. Purple circle represents uracil glycosylase inhibitor (UGI). E The adenine base editor (ABE). Light purple circle represents adenine deaminase TadA. F Prime editing technology. The prime editor (PE) is made up of a fusion protein of nCas9 (H840A) with reverse transcriptase and a prime editing guide RNA (pegRNA). G ZFNs, TALENs and CRISPR/Cas9 deliver double strand breaks (DSBs). DNA repair pathway includes the DNA non-homologous end joining (NHEJ) repair pathway and homology directed repair (HDR) pathway. DNA repair pathway produce different forms of genome modifications. H CBE generates base substitution of C•G to A•T without DSBs. I ABE make base substitution of A•T to G•C without DSBs. J PE generate precise genome modification of DNA substitution, insertion and deletion |
Table 1 Recent advances of CRISPR/Cas genome editing in economically important fruit crops |
Species | Promoter | Nuclease | Target Gene(s) | Target Trait(s) | Cas9 delivery method | Refence |
---|---|---|---|---|---|---|
Fragaria vesca | AtUBQ10 | Cas9 | TAA1, ARF8 | Fruit development | Agrobacterium-mediated | (Zhou et al., 2018) |
Fragaria vesca | 35S | Cas9 | YUCCA10 | Fruit development | Agrobacterium-mediated | (Feng et al., 2019) |
Fragaria vesca | 35S | Cas9 | SEP3 | Flower and fruit development | Agrobacterium-mediated | (Pi et al., 2021) |
Fragaria vesca | AtUBQ10 | Cas9 | AGL62, ALG80 | Fruit development | Agrobacterium-mediated | (Guo et al., 2022) |
Fragaria vesca | Ubi | nCas9 (D10A) | bZIPs1.1 | Fruit sugar content | Agrobacterium-mediated | (Xing et al., 2020) |
Fragaria vesca | Ubi | Cas9 | MYB10, CHS, PDS, UF3GT, F3H, LDOX | Fruit coloration | Agrobacterium-mediated | (Xing et al., 2018) |
Fragaria vesca | 35S | Cas9 | LAM | Plant architecture | Agrobacterium-mediated | (Feng et al., 2021a) |
Fragaria ananassa | 35S | Cas9 | TM6 | Flower and fruit development | Agrobacterium-mediated | (Martin-Pizarro et al., 2019) |
Fragaria ananassa | 35S | Cas9 | RAP | Fruit coloration | Agrobacterium-mediated | (Gao et al., 2020) |
Cucumis sativus | 35S | Cas9 | WIP1 | Flower development | Agrobacterium-mediated | (Hu et al., 2017) |
Cucumis sativus | 35S | Cas9 | SPT,ALC | Flower and fruit development | Agrobacterium-mediated | (Cheng et al., 2022) |
Cucumis sativus | 35S | Cas9 | HEC2 | Fruit development | Agrobacterium-mediated | (Wang et al., 2021e) |
Cucumis sativus | 35S | Cas9 | NS | Fruit development | Agrobacterium-mediated | (Liu et al., 2022) |
Citrullus lanatus | UBI | Cas9 | PSK1 | Resistance to Fusarium oxysporum | Agrobacterium-mediated | (Zhang et al., 2020b) |
Citrullus lanatus | 35S | Cas9 | WIP1 | Flower development | Agrobacterium-mediated | (Zhang et al., 2020a) |
Citrullus lanatus | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Tian et al., 2017) |
Citrullus lanatus | 35S | Cas9 | COMT1 | Fruit quality, abiotic stress | Agrobacterium-mediated | (Chang et al., 2021) |
Citrullus lanatus | 35S | Cas9 | NAC68 | Fruit sugar content | Agrobacterium-mediated | (Wang et al., 2021b) |
Citrullus lanatus | 35S | Cas9 | BG1 | Seed development | Agrobacterium-mediated | (Wang et al., 2021c) |
Cucumis melo | PcUbi4-2 | Cas9 | ROS1, CTR1-like | Fruit ripening | Agrobacterium-mediated | (Giordano et al., 2022) |
Vitis vinifera | 35S | Cas9 | IdnDH | Fruit quality | Agrobacterium-mediated | (Ren et al., 2016) |
Vitis vinifera | PcUbi4-2 | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Nakajima et al., 2017) |
Vitis vinifera | VvUBQ2 | Cas9 | TMT1, TMT2, PDS | Fruit sugar accumulation; Albino phenotype | Agrobacterium-mediated | (Ren et al., 2021) |
Vitis amurensis | 35S | Cas9 | PAT1 | Cold response | Agrobacterium-mediated | (Wang et al., 2021d) |
Vitis vinifera | 35S | Cas9 | CCD8 | Shoot branching | Agrobacterium-mediated | (Ren et al., 2020) |
Vitis vinifera | 35S | Cas9 | PR4b | Defense against the downy mildew | Agrobacterium-mediated | (Li et al., 2020a) |
Vitis vinifera | 35S | Cas9 | MLO3, MLO4 | Defense against the powdery mildew | Agrobacterium-mediated | (Wan et al., 2020) |
Vitis vinifera | 35S | Cas9 | AGL104 | Flower, fruit and seed development | Agrobacterium-mediated | (Sun et al., 2020b) |
Citrus sinensis | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Jia and Wang, 2014) |
Citrus sinensis | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Dutt et al., 2020) |
Citrus sinensis | 35S, Yao | LbCas12a | PDS, L0B1 | Resistant to citrus canker | Agrobacterium-mediated | (Jia et al., 2019) |
Citrus sinensis | 35S | Cas9 | NPR3 | Systemic acquired resistance (SAR) | Cationic lipid transfection with or without PEG | (Mahmoud et al., 2022) |
Citrus sinensis | CmYLCV | PC-ABE8e | LOB1 | Resistant to canker | Agrobacterium-mediated | (Huang et al., 2022) |
Citrus paradise | CmYLCV | nCas9 | ALS | resistant to the herbicide | Agrobacterium-mediated | (Huang et al., 2022) |
Malus prunifolia | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Nishitani et al., 2016) |
Malus domestica | PcUbi4-2 | nCas9 | ALS, PDS | Resistance to chlorsulfuron and albino | Agrobacterium-mediated | (Malabarba et al., 2020) |
Malus domestica | AtUBQ10 | Cas9 | DIPM4 | Resistance to fire blight disease | Agrobacterium-mediated | (Pompili et al., 2020) |
Malus domestica | AtUBQ10 | LbCas12a | PDS | Albino phenotype | Agrobacterium-mediated | (Schropfer and Flachowsky, 2021) |
Malus domestica | PcUbi4-2 | Cas9 | PDS, TFL1 | Albino phenotype; early flowering | Agrobacterium-mediated | (Charrier et al., 2019) |
Malus sieverii | ZmUbi, 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Zhang et al., 2021) |
Malus domestica | 35S | Cas9 | CNGC2 | Resistance to B. dothidea | Agrobacterium-mediated | (Zhou et al., 2020a) |
Malus domestica | 35S | Cas9 | MKK9 | Fruit color | Agrobacterium-mediated | (Sun et al., 2022) |
Solanum lycopersicum | 35S | Cas9 | SHR | Root development | Agrobacterium-mediated | (Ron et al., 2014) |
Solanum lycopersicum | 35S | Cas9 | AGO7 | Leaf morphology | Agrobacterium-mediated | (Brooks et al., 2014) |
Solanum lycopersicum | 35S, AtUBQ | Cas9 | PDS, PIF4 | Albino phenotype, light signal transduction | Agrobacterium-mediated | (Pan et al., 2016) |
Solanum lycopersicum | 35S | Cas9 | ARF4 | Plant growth, resistance to abiotic stress | Agrobacterium-mediated | (Bouzroud et al., 2020) |
Solanum lycopersicum | 35S | Cas9 | SRM1-like | Leaf development | Agrobacterium-mediated | (Tang et al., 2022) |
Solanum lycopersicum | 35S | nCas9 (D10A) | DELLA, ETR1 | Hormone signaling | Agrobacterium-mediated | (Shimatani et al., 2017) |
Solanum lycopersicum | 35S | Cas9 | SP, SP5G, ER | Plant architecture | Agrobacterium-mediated | (Kwon et al., 2020) |
Solanum lycopersicum | 35S | Cas9 | BOP | Inflorescence development | Agrobacterium-mediated | (Xu et al., 2016) |
Solanum lycopersicum | 35S | Cas9 | DOF9 | Inflorescence and flower development | Agrobacterium-mediated | (Hu et al., 2022) |
Solanum lycopersicum | Ubi | Cas9 | ORRM4 | Fruit ripening | Agrobacterium-mediated | (Yang et al., 2017) |
Solanum lycopersicum | Ubi | Cas9 | LncRNA1459 | Fruit ripening | Agrobacterium-mediated | (Li et al., 2018a) |
Solanum lycopersicum | 35S, PcUbi4-2 | Cas9 | IAA9 | Fruit development | Agrobacterium-mediated | (Ueta et al., 2017) |
Solanum lycopersicum | PcUbi4-2 | Cas9 | GAD2, GAD3 | Fruit quality | Agrobacterium-mediated | (Nonaka et al., 2017) |
Solanum lycopersicum | 35S | Cas9 | PSY1 | Fruit color | Agrobacterium-mediated | (Filler Hayut et al., 2017) |
Solanum lycopersicum | Ubi | Cas9 | MIR164A | Fruit ripening and chloroplast development | Agrobacterium-mediated | (Lin et al., 2022b) |
Solanum lycopersicum | PcUbi4-2 | Cas9 | KIX9; SlKIX8 | Plant organ size | Agrobacterium-mediated | (Swinnen et al., 2022) |
Solanum lycopersicum | 35S | Cas9 | CRCa | Floral meristem determinacy | Agrobacterium-mediated | (Castaneda et al., 2022) |
Solanum lycopersicum | 35S | Cas9 | CLV3, WOX9, TFL1, | Floral organ number, fruit size | Agrobacterium-mediated | (Rodriguez-Leal et al., 2017) |
Solanum lycopersicum | 35S | Cas9 | ENO | Fruit size | Agrobacterium-mediated | (Yuste-Lisbona et al., 2020) |
Solanum lycopersicum | Ubi | Cas9 | MYC2 | Plant development, disease resistance | Agrobacterium-mediated | (Shu et al., 2020) |
Solanum lycopersicum | 35S | Cas9 | AGL6 | Fruit development | Agrobacterium-mediated | (Klap et al., 2017) |
Solanum lycopersicum | 35S | Cas9 | GGP1 | Fruit quality | Agrobacterium-mediated | (Deslous et al., 2021) |
Solanum lycopersicum | 35S | Cas9 | INVINH1,VPE5 | Fruit sugar content | Agrobacterium-mediated | (Wang et al., 2021a) |
Musa acuminata | 35S | Cas9 | RAS-PDS | Albino phenotype | Agrobacterium-mediated | (Kaur et al., 2018) |
Musa acuminata | ZmUbi1, 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Naim et al., 2018) |
Musa acuminata | Ubi | Cas9, LbCas12a | PDS | Albino phenotype | PEG mediated RNP | (Wu et al., 2020) |
Musa acuminata | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium-mediated | (Ntui et al., 2020) |
Musa acuminata | 35S | Cas9 | LCYε | Fruit quality | Agrobacterium-mediated | (Kaur et al., 2020) |
Musa acuminata | Ubi | Cas9 | GA20OX2 | Plant height | Agrobacterium-mediated | (Shao et al., 2020) |
Musa acuminata | Ubi | Cas9 | ACO1 | Ethylene production; Fruit shelf | Agrobacterium-mediated | (Hu et al., 2021) |
Musa acuminata | Ubi | Cas9 | PDS | Albino phenotype | Protoplast transformation | (Zhang et al., 2022) |
Musa acuminata | 35S | Cas9 | CCD4 | Carotenoids metabolism | Particle bombardment | (Awasthi et al., 2022) |
Actinidia chinensis | 35S | Cas9 | PDS | Albino phenotype | Agrobacterium method | (Wang et al., 2018b) |
Actinidia chinensis | 35S | Cas9 | FLCL | Flower development | Agrobacterium method | (Voogd et al., 2022) |
Actinidia chinensis | 35S | Cas9 | SyGl, CEN4 | Flower development | Agrobacterium method | (Varkonyi-Gasic et al., 2021) |
Actinidia chinensis | Ubi, 35S | Cas9 | CEN, CEN4 | Plant stature, early flowering | Agrobacterium method | (Varkonyi-Gasic, et al., 2019) |
Application of genome editing in fruit crops
Plant development
Non-climacteric fruits (strawberry, grape, watermelon, cucumber, citrus)
Fig. 3 Strawberry fruit structure and a model illustrating the regulatory pathway during fruit initiation and growth. A Double fertilization promotes biosynthesis of auxin and GA in the seed. Auxin and GA can stimulate receptacle development after being transported to the receptacle. B A diagram of receptacle illustrating the regulatory mechanism of fruit set. FveRGA1 is shown as a central player. The red lines indicate regulatory actions post-fertilization. Positive (arrows) or negative (bar) regulations are indicated. Adapted from (Feng et al. 2019; Zhou et al. 2021; Guo et al. 2022) |
Climacteric fruits (apple, tomato, banana, kiwifruit)
Fig. 4 The interaction among ABA, auxin, and GA in regulating strawberry and tomato fruit development. A A diagram of strawberry fruit illustrating the regulatory mechanism during fruit ripening. In the achene, the expression of ABA biosynthesis gene (FaNCED6) increases, and the expression of ABA metabolism gene (FaCYP707A4, FaCYP722A) decreases, which resulted in a high ABA level. On the other hand, the expression of FaPIN2/3/5 and FaYUC1/2/10/11 decreases, which resulted in a reduced auxin and GA level. The feedforward loop of ABA level was activated, and the ripening related transcription factors (MYB79, MYB10) were activated to ensure fruit firmness and anthocyanin biosynthesis. As fruit ripens, JA level accumulates to ensure flavor formation. B In tomato, seeds are the site of auxin production. Auxin was also transported to the surrounding tissues to stimulate fruit growth. However, during fruit ripening, SlEIN3 and SlTAGL1, activate ethylene production and form a positive feedback circuit. SlEIN3 and SlTAGL1 form a complex and promote fruit ripening by regulating transcription factors SlCRTISO, SlPL, SlEXP1, SlCEL2 etc. On the other hand, the expression of SlCRTISO, SlPL, SlEXP1 and SlCEL2 is also related to ethylene production. However, whether there is a direct regulation is not clear. Adapted from (Kang et al. 2013a; Liao et al. 2018; Cao et al. 2020; Chen et al. 2020; Li et al. 2022) |
Plant immunity
Biotic stresses
Abiotic stresses
Challenges and improvements of applying genome editing in fruit crops
Improving genome editing efficiency by optimizing the expression of sgRNA and Cas proteins
Improving the delivery efficiency of CRISPR/Cas reagents
Improving the specificity of genome editing
Optimizing transformation and regeneration systems
Obtaining the transgene-free plants
Future directions and remarks
Application of novel techniques in fruit crops
Application of novel CRISPR/Cas-derived transcriptional activation/ repression and epigenetic regulation platforms
Fig. 5 CRISPR/dCas9-based epigenetic modifier during fruit ripening process in tomato, sweet orange, and strawberry. A CRISPR/dCas9 based epigenetic modifier in tomato fruit ripening. The DNA demethylase can be engineered to fuse to dCas9 system. In tomato fruit ripening process, the expression of DNA demethylase DME-LIKE 2 (DML2) increases, leading to decrease of 5-methlcytosine (5mC) DNA level at various gene loci, such as RIN, CNR, and NOR. B CRISPR/dCas9 based epigenetic modifier in sweet orange fruit ripening process. DNA methyltransferase could be engineered to fuse to dCas9 system. During sweet orange fruit ripening, DNA demethylase genes are downregulated, leading to the upregulation of 5mC DNA methylation level. C CRISPR/dCas9 based epigenetic modifier in strawberry fruit ripening process. DNA methyltransferase could be engineered to fuse to dCas9 system for promoting strawberry fruit ripening. As strawberry fruit ripening, the activity of RNA-directed DNA methylation is reduced, resulting in a decrease in the DNA methylation level. Diagrams were drawn based on publications (Liu et al. 2015; Cheng et al. 2018; Huang et al. 2019; Chen et al. 2020) |