Multiplex CRISPR-Cas9 Gene-Editing Can Deliver Potato Cultivars with Reduced Browning and Acrylamide

Abstract

Storing potato tubers at cold temperatures, either for transport or continuity of supply, is associated with the conversion of sucrose to reducing sugars. When cold-stored cut tubers are processed at high temperatures, with endogenous asparagine, acrylamide is formed. Acrylamide is classified as a carcinogen. Potato processors prefer cultivars which accumulate fewer reducing sugars and thus less acrylamide on processing, and suitable processing cultivars may not be available. We used CRISPR-Cas9 to disrupt the genes encoding vacuolar invertase (VInv) and asparagine synthetase 1 (AS1) of cultivars Atlantic and Desiree to reduce the accumulation of reducing sugars and the production of asparagine after cold storage. Three of the four guide RNAs employed induced mutation frequencies of 17–98%, which resulted in deletions, insertions and substitutions at the targeted gene sites. Eight of ten edited events had mutations in at least one allele of both genes; for two, only the VInv was edited. No wild-type allele was detected in both genes of events DSpco7, DSpFN4 and DSpco12, suggesting full allelic mutations. Tubers of two Atlantic and two Desiree events had reduced fructose and glucose concentrations after cold storage. Crisps from these and four other Desiree events were lighter in colour and included those with 85% less acrylamide. These results demonstrate that multiplex CRISPR-Cas9 technology can generate improved potato cultivars for healthier processed potato products.

Reducing Acrylamide Formation Potential by Targeting Free Asparagine Accumulation in Seeds

“…More potential strategies can include optimal harvest time and managing storage condition irrigation and fertilization, all of which can affect overall sugar levels in crops . However, it is worth noting that reducing sugars are precursors to many Maillard products that have favorable attributes, such as browning, aroma, and flavor; thus, a decrease in the sugar content could be detrimental to product quality.…”

“…Reducing Asn accumulation in cereals is considered an effective strategy because Asn was shown to be a limiting factor in acrylamide formation. 14 Asn is synthesized via three routes: 20 the first is the conversion of oxosuccinamic acid (the oxo analogue of Asn) to Asn by asparagine−oxo-acid transaminase; the second is the conversion of β-cyanoalanine to Asn catalyzed by βcyanoalanine synthase; and the third is the conversion of aspartate to Asn catalyzed by Asn synthetase (AsnS). The βcyanoalanine synthase route occurs only in the presence of cyanide, and the Asn−oxo-acid transaminase route is marginal in plants; 23 thus, the Asn synthetase pathway is the most significant route.…”

New Frontiers in Potato Breeding: Tinkering with Reproductive Genes and Apomixis

“…In potatoes, substantial advancements have been realized through genome editing [ 190 ], particularly in enhancing traits related to tuber quality. Notable achievements encompass the modification of starch content [ 191 ], increased resistance to bruising (resulting in reduced enzymatic browning) [ 189 ], and enhanced tolerance to cold storage achieved through the targeted knockout of the Vacuolar Invertase gene ( VInv ) [ 192 ].…”

“…In recent years, significant advances in gene-editing technology have expanded the possibilities for editing multiple genes simultaneously, whether targeting a single complex trait or multiple distinct traits. For instance, a study by Ly et al [ 192 ] successfully edited the VInv and Asparagine Synthetase 1 ( AS1 ) genes associated with acrylamide production. Massa et al [ 195 ] simultaneously knocked out Polyphenol Oxidases ( StPPO s) and VInv genes in cultivars Spunta and Atlantic to enhance industrial and nutritional potato quality.…”

Genetic architecture of tuber-bound free amino acids in potato and effect of growing environment on the amino acid content

“…Despite the need, polyploidy and high heterozygosity make breeding and selection of low acrylamide-forming potential potato cultivars for healthy fries and chips challenging. Possibilities of using the Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) system to manipulate reducing sugars and aspargine metabolism in potatoes are already being considered 29 . Another amino acid affecting flavor is methionine, a precursor of methional, a major volatile associated with the aroma of potato chips and french fries 30 .…”