CRISPR / Cas to improve the nutritional and industrial quality of potatoes

MASSA GABRIELA ALEJANDRA

Simposio; X ENCUENTRO LATINOAMERICANO Y DEL CARIBE DE BIOTECNOLOGÍA AGROPECUARIA Y XI SIMPOSIO REDBIO ARGENTINA; 2019

Potato isone of the most important foods in human dietary. Cultivatedpotato has a tetraploid, highly heterozygous genome and it is clonallypropagated. These characteristics make conventional breeding and geneticanalysis very challenging. In this context, the CRISPR/Cas9 technology has beensuccessfully utilized in potato for precise Genetic Edition. Two fundamental aspects of the nutritional and industrialquality in potatoes are the behavior of the varieties respect to"cold-induced sweetening" and "enzymatic browning" intuber. Cold-induced sweetening occurs when harvested tubers are stored attemperatures below 10 °C and for the action of vacuolar invertase the tuber accumulatereducing sugars. The enzymatic browning is for the action of Polyphenoloxidases (PPO) enzymes that catalyze the oxidation of phenolic compounds in potatoesproducing black spots on the tubers, with the consequent economic losses due todiscarding and reduction in consumer acceptance. Our objective is todevelopment varieties with reduction of cold-induced sweetening and enzymaticbrowning by CRISPR/Cas9 system. In a first approximation we obtained constructions withtwo RNA guide specific for vacuolar invertase (Vac-Inv) and polyphenol oxidase2 (StPPO2) genes. A Golden Gate assembled binary vector which codifiesfor the CRISPR/Cas9 reagents, was stable integrated in potato genome via Agrobacterium tumefaciens. The CRISPRfor Vac-Inv was evaluated by protoplast transfection previously totransformation. We analyzed the transgenic plants by HRFA and CAPS and weobtained 9,7 % of edited plants in at least one allele of StPPO2 gene. Otherobjective is to obtain edited potatoes plants in both genes by protoplasttransfections with ribonucleoprotein (gRNA/Cas9), DNA-free method. The GErepresents the most promising and safe approach to achieve a reduction inproduction costs, a decrease in the discarding of raw material and a product ofhigher nutritional quality for the health of consumers.