TABLE 1.
List of recent successes in the improvement of crops with the use of clustered regularly interspaced short palindromic repeats (CRISPR-Cas).
| Associated trait | Target gene/Cas variant | Gene function | Modification efficiency/Result | Cultivar | Transformation method | References |
|---|---|---|---|---|---|---|
| Yield (seed size and seed number) | BnaEOD3 | Controls seed size, silique length, and seed production in rapeseed | Knockout | Brassica napus | Agrobacterium-mediated transformation | Khan et al. (2020) |
| Cas variant: Cas9 | Seed weight increased in the quadruple mutants by 13.9% | |||||
| Yield (pod shatter-resistant) | BnSHP1 | BnSHP1A09 regulation of lignin composition at the dehiscence zone | Knockout | Brassica napus | Agrobacterium-mediated transformation | Zaman et al. (2021) |
| BnSHP2 | Pod-shattering resistance index (SRI) increased in mutant lines (0.31) compared to the wild-type (WT) (0.036) | |||||
| Cas variant: Cas9 | ||||||
| Yield (plant architecture) | BnaBP | Regulation of pedicel bending and leaf morphogenesis in Arabidopsis | Knockout | Brassica napus | Agrobacterium-mediated transformation | Fan et al. (2021) |
| Cas variant: Cas9 | Downregulation of BnaBP genes decreases the branch angle to create more compact plants | Plant height decreased in the mutant plants by 15.8%–16.9% | ||||
| Branch angle decreased from 84° (WT) to 14° in mutant plants | ||||||
| Plant yield and architecture | BnaMAX1 | bnaMAX1 gene controls axillary bud outgrowth and plant height | Knockout | Brassica napus | Agrobacterium tumefaciens mediated floral dip method | Zheng et al. (2020) |
| Cas variant: Cas9 | Plant height decreased by 31.9%–36.5% and total silique number increased by 62.3%–71.8% in mutant plants compared to the WT | |||||
| Fatty acid composition and oil content (oil content) | BnLPAT2 | BnLPAT2/BnLPAT5 plays a major role in regulating the morphology and number of oil bodies | Knockout | Brassica napus | Agrobacterium-mediated hypocotyl transformation method | Zhang et al. (2019) |
| BnLPAT5 | Mutation frequency: 17%–68% | |||||
| Cas variant: Cas9 | No mutation observed in off-target sites | |||||
| Fatty acid composition and oil content (oleic acid content) | BnaFAD2 | Fatty acid desaturase-2 (FAD2) gene impacts fatty acids, mainly oleic, linoleic, and linolenic, in oilseed plants | 74.1% short-nucleotide alterations (≤3 bp), with 51.9% single nucleotide insertions and deletions | Brassica napus | Agrobacterium-mediated hypocotyl transformation method | Huang et al. (2020) |
| Cas variant: Cas9 | ||||||
| Isoflavone content | GmF3H1, GmF3H2 and GmFNSII-1 | Isoflavone synthesis | Knockout | Glycine max | A. rhizogenes-mediated method and Agrobacterium-mediated cot node transformation | Zhang et al. (2020) |
| Cas variant: Cas9 | Mutation efficiency increased by 44.44%, and triple gene mutation observed in T0 transgenic plants | |||||
| Yield (blocking OsAAP3 increases grain yield by increasing tiller number in rice) | OsAAP3 | OsAAP3 gene increases rice tiller number and elongation of outgrowth bud by regulating the concentrations of Lys, His, Ala, Asp, Arg, Gln, Gly, Tyr, and Thr in rice | Knockout and overexpression | Oryza sativa | Agrobacterium-mediated transformation | Lu et al. (2018) |
| Cas variant: Cas9 | Tiller numbers increased in OsAAP3 knockout lines and decreased in OsAAP3 OE lines compared to WT plants | |||||
| Yield (grain weight, grain number, and grain size) | OsGS3, OsGW2 and OsGn1a | Negatively regulate grain weight, width, number, and size | Off-target mutation Mutation frequency: 66.7%–100% |
Oryza sativa | Agrobacterium-mediated transformation | Zhou et al. (2019) |
| Cas variant: Cas9 | GN1a encodes a cytokinin oxidase/dehydrogenase, OsCKX2 that negatively regulates grain size | |||||
| Increased resistance to bacterial blight | OsSWEET11 (PthXo1) and OsSWEET14 (PthXo3/AvrXa7) | SWEET genes are susceptibility (S) genes and encode sugar transporter proteins. As a result, recessive alleles of these SWEET genes confer resistance | Knockout | Oryza sativa | Agrobacterium-mediated transformation | Xu et al. (2019) |
| Cas variant: Cas9 | Using the CRISPR/Cas9 system, a new germplasm named MS134K was developed with mutated EBE alleles of OsSWEET13, OsSWEET14, and OsSWEET11, providing excellent resistance to Xoo strains | |||||
| Cotton genome editing efficiency by Cas 12 | Deoxyglucose-5-phosphate synthase (GhCLA) | Terpenoid biosynthesis | Knockout | Gossypium hirsutum | Agrobacterium-mediated genetic transformation | Wang et al. (2020) |
| Cas variant: Cas12b (C2c1) from Alicyclobacillus acidoterrestris (AacCas12b) | Mutation rate: ∼20% | |||||
| Yield (disease resistance) | xopV | XopV suppresses PTI peptidoglycan-triggered response in rice | Knockout | Oryza sativa | Electro-transformation | Yan et al. (2023) |
| Cas variant: Cas12a from Francisella novicida | CRISPR-NHEJ method produced high editing frequencies (40.91%–95.45%) | |||||
| Plant architecture and fruit ripening | SlBRI1, SlRIN | slBRI1 gene performs an essential function in controlling plant architecture | Knockout | Solanum lycopersicum | Agrobacterium-mediated genetic transformation | Niu et al. (2020) |
| Cas variant: XNG-Cas9 | SlRIN gene has a vital role in the ripening of fruits | XNG-Cas9 edited the AGG site with 13.3% efficiency. Other editing efficiencies: 23.5% at GGC, 22.2% at GGT, and 15.2% at TGA sites., Mutation efficiency of 15.8% at AGA site | ||||
| Carotenoid biosynthesis | PDS3 (phytoene desaturase) | PDS3 gene positively regulates carotenoid biosynthesis | Knockout | Arabidopsis thaliana | Protoplast transfection | Pausch et al. (2020) |
| Cas variant: Cas Ф | 8–10 bp deletions in PDS3 genes | |||||
| Fatty acid composition and oil content (oil content) | Glyma10g42470 and FAD2-A | Controls oleic acid content in developing soybean seed | Knockout | Glycine max |
Agrobacterium-mediated soybean hairy root transformation Agrobacterium rhizogenes |
Duan et al. (2021) |
| Cas variant: Cpf1 from Lachnospiraceae bacterium ND 2006 (LbCpf1) | CRISPR/LbCpf1 induced deletions of huge chromosomal segments with up to 91.7% editing efficiency | |||||
| Ureide biosynthesis | XDH, NSH1, NSH2, XMPP and GSDA | All genes mentioned are involved in ureide biosynthesis | The onset of hairy root development activates the automatic repair mechanism when Cas9 induces double-strand breaks. Determined the xanthosine and guanosine are key metabolites needed for ureide production | Phaseolus vulgaris | R. rhizogenes | Voß et al. (2022) |
| Cas variant: Cas9 | ||||||
| Loss of seed and pod development | Vu-SPO11 | Mutation in this gene causes fertility loss, with no seed or pod development | Knockout | Vigna unguiculata | Agrobacterium-mediated genetic transformation) | Che et al. (2021) |
| Cas variant: spCas9 | Effective gene editing | |||||
| Modification frequencies: 4%–37% | ||||||
| Disease resistance | Two CsWRKY22 alleles | Decreases Wanjincheng orange susceptibility to Xanthomonas citri subsp. Citri | Indels and nucleotide substitutions | Citrus sinensis | Agrobacterium-mediated genetic transformation | Wang et al. (2019) |
| Mutation rates of mutant lines: | ||||||
| Cas variant: Cas9 | W1-1: 85.7% | |||||
| W2-2: 79.2% | ||||||
| W2-3: 68.2% | ||||||
| Crop improvement | OsALS, NRT1.1B, OsCDC48and OsWaxy | OsALS: provides imazamox herbicide resistance in rice | Base edit | Oryza sativa | Agrobacterium-mediated genetic transformation | Xu et al. (2019) |
| NRT1.1B gene increases nitrogen use efficiency in rice plants | At the various targets, the editing efficiency increased 2- to 3-fold in the three high-fidelity Cas9 variants | |||||
| Cas variant: Three SpCas9 variants, SpCas9-HF2, HypaCas9 and eSpCas9 (1.1) | OsCDC48 regulates cell death and senescence | |||||
| OsWaxy plays a vital role in granule-bound starch biosynthesis | ||||||
| Crop improvement (chlorsulfuron-resistant plants) | SlALS1 | Acetolactate synthase (ALS) gene is important for branched-chain amino acid biosynthesis | Base edit | Solanum tuberosum and Solanum lycopersicum | Agrobacterium-mediated genetic transformation | Veillet et al. (2019) |
| Cas variant: nCas9 cytidine base editing system | Homologous gene selection and sequencing | |||||
| Modification efficiency: 71% |