CRISPR-Cas Gene Editing Technology in Potato

Abstract

Potato (Solanum tuberosum L.) is one of the most important food crops in the world, ranking fourth after rice, maize, and wheat. Potatoes are exposed to biotic and abiotic environmental factors, which lead to economic losses and increase the possibility of food security threats in many countries. Traditional potato breeding faces several challenges, primarily due to its genetic complexity and the time-consuming nature of the process. Therefore, gene editing-CRISPR-Cas technology-allows for more precise and rapid changes to the potato genome, which can speed up the breeding process and lead to more effective varieties. In this review, we consider CRISPR-Cas technology as a potential tool for plant breeding strategies to ensure global food security. This review summarizes in detail current and potential technological breakthroughs that open new opportunities for the use of CRISPR-Cas technology for potato breeding, as well as for increasing resistance to abiotic and biotic stresses, and improving potato tuber quality. In addition, the review discusses the challenges and future perspectives of the CRISPR-Cas system in the prospects of the development of potato production and the regulation of gene-edited crops in different countries around the world.

Keywords: CRISPR-Cas; abiotic stress; biotic stress; disease resistance; drought; gene editing; improve breeding process; potato; tuber quality; viral resistance.

Figure 1

Conventional plant breeding (left panel) and CRISPR-Cas editing technology (right panel) for the potato breeding process. Conventional plant breeding (left panel): (1) Year 1—cross parents: selection of parental varieties, hybridization, berry formation, and true potato seed (TPS) extraction; (2) Year 2—seedling stage: the true potato seed grown as a seedling and the production of seedling tubers; (3) initial evaluation trails: Year 3—A-Clones (F₁C₁), grown from seedling tubers, grown at 2 locations on unreplicated plots; Year 4—B-Clones (F₁C₂), grown on at least 3 locations (replicated plots); Year 5—C-Clones (F₁C₃), grown on at least 5 locations (replicated plots); from Year 3 to Year 5 (F₁C₁-F₁C₃)—tuber morphology evaluation: tuber color and shape, quality, and resistance to biotic and abiotic stresses by Marker Assisted Selection (MAS); (4) advanced stage trails: Year 6—D-Clones (F₁C₄), grown on 5 or more locations (replicated plots); Year 7—E-Clones (F₁C₅), grown on 5 or more locations (replicated plots); from Year 5 and 6 (F₁C₄–F₁C₅)—phenotypic evaluation, yield, quality, and disease resistance; (5) official testing: Year 8–9 (F₁C₆–F₁C₇)—selection, distinctness, uniformity, and stability (DUS) testing, DNA fingerprinting, and the release of an advanced potato clone; and (6) introduction and multiplication: Year 10—commercialization of an advanced potato clone. Conventional potato breeding using MAS may take 10 years to develop a potato variety. CRISPR-mediated GE (right panel): (1) definition of the target gene and design of sgRNA (Year 1, 1 week); (2) assembly of the CRISPR/Cas9 vector (Year 1, 1–3 weeks); (3) transformation of potato cells and the obtaining and screening of mutated plants (T₀) (Year 1, 1 month); (4) obtaining gene-edited plants lacking vector sequences in T₁–T₂ generations (Year 1, 10 months), and screening of CRISPR-Cas mutated plants using Sanger sequencing, CAPS analysis, or T7 assays, etc; (5) phenotypic evaluation—after confirming gene edits in the plants, phenotypic evaluation stage proceeds as in conventional breeding to evaluate yield, quality, and disease resistance (Year 2); (6) official testing: Year 3–4—selection, DUS testing, DNA fingerprinting, and the release of the edited potato variety; and (7) introduction and multiplication: Year 5—commercialization of the edited potato variety. The development of a potato variety may take 5 years; however, crop improvement using CRISPR-Cas technology is limited due to its cost, low efficiency, and regulatory issues.

Figure 2

Using CRISPR-Cas technology to improve potato resistance to biotic stresses. Biotic stress includes fungal, viral, and bacterial diseases. In wild type potato (left panel) StNRL1, StDND1, StCHL1, StDMR6-1, StPM1, StCCoAOMT, StERF3, StDMP2, StMC7, and StSR4 genes negatively regulated late blight resistance, while StDMR6-1 negatively regulated late blight, early blight, and common scab; StCoilin, SteIF4E, SteIF4E1, StP3, StCI, StNib, StCP, StPI, StHC-Pro, StCl1, StCl2, and StVPg genes negatively regulated resistance to potato virus Y, while CP multiplex virus resistance PVY, PVS, PVX, or PLRV; NPR3 gene negatively regulated resistance to Clso. In CRISPR-Cas edited plants (right panel) knockout of susceptibility (S) genes (ΔStNRL1, ΔStDND1, ΔStCHL1, ΔStDMR6-1, ΔStPM1, ΔStCCoAOMT, ΔStERF3, ΔStDMP2, ΔStMC7, ΔStSR4, ΔStCoilin, ΔSteIF4E, ΔSteIF4E1, ΔStNPR3) and virus RNA targeting (ΔStP3, ΔStCI, ΔStNib, ΔStCP, ΔStPI, ΔStHC-Pro, ΔStCl1, ΔStCl2, ΔStVPg) improved resistance to biotic stresses. The loss of function genes ΔStNRL1, ΔStDND1, ΔStCHL1, ΔStDMR6-1, ΔStPM1, ΔStCCoAOMT, ΔStERF3, ΔStDMP2, ΔStMC7, and ΔStSR4 improve resistance to P. infestans. The loss-of-function gene ΔStNPR3 improves resistance to Clso. The loss-of-function genes ΔStCoilin, ΔSteIF4E, ΔSteIF4E1, ΔStP3, ΔStCI, ΔStNib, ΔStCP, ΔStPI, ΔStHC-Pro, ΔStCl1, ΔStCl2, and ΔStVPg improve resistance to potato virus Y. The ΔStCP gene improves multiplex virus resistance to PVY, PVS, PVX, and/or PLRV.

Figure 3

Using CRISPR-Cas technology to improve potato tolerance to abiotic stresses. Abiotic stresses include drought, salinity, high-light, Pi transportation, and iron regulation. In wild type potato (left panel), genes involved in negative regulation include the following: drought (StDMR6-1, StDRO2); high-light (StAOX); salt stress (StDMR6-1); Pi transportation (StMYB44), and iron regulation (StbHLH47). In CRISPR-Cas edited plants (right panel), knockout of target genes for drought (ΔStDMR6-1, ΔStDRO2), high-light (ΔStAOX), salt stress (ΔStDMR6-1), Pi transportation (ΔStMYB44), and iron regulation (ΔStbHLH47) improved resistance to abiotic stresses. The loss of function of the following genes: ΔStDMR6-1—improved drought and salt tolerance; ΔStDRO2 and ΔStAOX—high-light tolerance; ΔStMYBb44—improved Pi transportation; and ΔStbHLH47—iron regulation. Nonsense mutation of ΔStDRO2 improved drought tolerance.

Figure 4

Using CRISPR-Cas gene editing technology for enhancement of potato qualities. In CRISPR-Cas edited plants, knockout of target genes, StSN2 StCDF1, StSP6A and StIT1, enhanced potato tuberization; StF3H gene was involved in tuber skin color change, from red to yellow; StPPO, StPPO1, and StPPO2 genes reduced enzymatic browning; StFtsZ1, StSS5, and StSS6 genes altered starch granule size; StGWD1 gene improved starch granule; StSSR2 and St16DOX genes reduced level of SGA; StGBSS, StGBSSI, StSBE1, StSBE2, and StSBE3 genes regulated amylopectin/amylose ratio; StMYB210 gene stopped anthocyanin synthesis; StVinv and StAS1 genes reduced sugars and low acrylamide levels; StVinv gene reduced fructose and glucose concentrations after cold storage; StInvVac gene was responsible for long-term cold storage and bruising resistance; StABCG1 gene decreased suberin production.

Figure 5

Using CRISPR-Cas gene editing technology to improve starch quality. The starch biosynthetic pathway is shown (left panel). In wild type potato (right upper panel), the StFtsZ1 gene is involved in starch granule size; the StSS5 and StSS6 genes are involved in starch granule formation; the StGWD1 gene catalyzes the phosphorylation of the glucose chain of starch; the StGBSS and StGBSSI genes are responsible for amylose content; the StSBE1, StSBE2, and StSBE3 genes regulate amylose content. In CRISPR-Cas edited plants (right lower panel), knockout of target genes included the following: ΔStFtsZ1 gene altered starch granule size; ΔStSS5 and ΔStSS6 genes increased formation of multiple starch granules; ΔStGWD1 reduced phosphorylation; ΔStGBSS and ΔStGBSSI decreased amylose content; and ΔStSBE1, ΔStSBE2, and ΔStSBE3 genes increased amylopectin content.