An alternative pathway to plant cold tolerance in the absence of vacuolar invertase activity.

To cope with cold stress, plants have developed antioxidation strategies combined with osmoprotection by sugars. In potato (Solanum tuberosum) tubers, which are swollen stems, exposure to cold stress induces starch degradation and sucrose synthesis. Vacuolar acid invertase (VInv) activity is a significant part of the cold-induced sweetening (CIS) response, by rapidly cleaving sucrose into hexoses and increasing osmoprotection. To discover alternative plant tissue pathways for coping with cold stress, we produced VInv-knockout lines in two cultivars. Genome editing of VInv in 'Désirée' and 'Brooke' was done using stable and transient expression of CRISPR/Cas9 components, respectively. After storage at 4°C, sugar analysis indicated that the knockout lines showed low levels of CIS and maintained low acid invertase activity in storage. Surprisingly, the tuber parenchyma of vinv lines exhibited significantly reduced lipid peroxidation and reduced H2 O2 levels. Furthermore, whole plants of vinv lines exposed to cold stress without irrigation showed normal vigor, in contrast to WT plants, which wilted. Transcriptome analysis of vinv lines revealed upregulation of an osmoprotectant pathway and ethylene-related genes during cold temperature exposure. Accordingly, higher expression of antioxidant-related genes was detected after exposure to short and long cold storage. Sugar measurements showed an elevation of an alternative pathway in the absence of VInv activity, raising the raffinose pathway with increasing levels of myo-inositol content as a cold tolerance response.

Chilling induces sugar and ABA accumulation that antagonistically signals for symplastic connection of dormant potato buds.

Endodormancy (ED) is a crucial stage in the life cycle of many perennial plants. ED release requires accumulating a certain amount of cold exposure, measured as chilling units. However, the mechanism governing the effect of chilling on ED duration is poorly understood. We used the potato tuber model to investigate the response to chilling as associated with ED release. We measured the accumulation of specific sugars during and after chilling, defined as sugar units. We discovered that ED duration correlated better with sugar units accumulation than chilling units. A logistic function was developed based on sugar units measurements to predict ED duration. Knockout or overexpression of the vacuolar invertase gene (StVInv) unexpectedly modified sugar units levels and extended or shortened ED, respectively. Silencing the energy sensor SNF1-related protein kinase 1, induced higher sugar units accumulation and shorter ED. Sugar units accumulation induced by chilling or transgenic lines reduced plasmodesmal (PD) closure in the dormant bud meristem. Chilling or knockout of abscisic acid (ABA) 8'-hydroxylase induced ABA accumulation, in parallel to sweetening, and antagonistically promoted PD closure. Our results suggest that chilling induce sugar units and ABA accumulation, resulting in antagonistic signals for symplastic connection of the dormant bud.

[Genotype-Specific Features of Cold-Induced Sweetening Process Regulation in Potato Varieties Nikulinsky, Symfonia, and Nevsky].

In this study, we performed expression analysis of genes associated with cold-induced sweetening in potato tubers: vacuolar invertase (Pain-1), sucrose synthase (SUS4), and invertase inhibitor (InvInh2). Potato varieties Nikulinsky, Symfonia, and Nevsky were used. All three varieties were found to accumulate sugars at low temperatures; the maximum accumulation of reducing sugars was observed at 4°C. It was found that the expression pattern of genes associated with cold-induced sweetening differs depending on the variety and storage duration. The increased expression of vacuolar invertase and its inhibitor is more pronounced at the beginning of storage period, whereas the increased expression of sucrose synthase is more pronounced after 3 months of storage. At early storage periods, high expression of invertase and low expression of inhibitor is observed in the Dutch variety Symfonia, and vice versa in the Russian varieties Nikulinsky and Nevsky. The involvement of the studied genes in the process of cold-induced sweetening is discussed.

CRISPR/Cas-mediated knockdown of vacuolar invertase gene expression lowers the cold-induced sweetening in potatoes.

MAIN CONCLUSION: VInv gene editing in potato using CRISPR/Cas9 resulted in knockdown of expression and a lower VInv enzymatic activity resulting in a decrease in post-harvest cold-storage sugars formation and sweetening in potatoes. CRISPR-Cas9-mediated knockdown of vacuolar invertase (VInv) gene was carried out using two sgRNAs in local cultivar of potato plants. The transformation efficiency of potatoes was found to be 11.7%. The primary transformants were screened through PCR, Sanger sequencing, digital PCR, and ELISA. The overall editing efficacy was determined to be 25.6% as per TIDE analysis. The amplicon sequencing data showed maximum indel frequency for potato plant T12 (14.3%) resulting in 6.2% gene knockout and 6% frame shift. While for plant B4, the maximum indel frequency of 2.0% was found which resulted in 4.4% knockout and 4% frameshift as analyzed by Geneious. The qRT-PCR data revealed that mRNA expression of VInv gene was reduced 90-99-fold in edited potato plants when compared to the non-edited control potato plant. Following cold storage, chips analysis of potatoes proved B4 and T12 as best lines. Reducing sugars' analysis by titration method determined fivefold reduction in percentage of reducing sugars in tubers of B4 transgenic lines as compared to the control. Physiologically genome-edited potatoes behaved like their conventional counterpart. This is first successful report of knockdown of potato VInv gene in Pakistan that addressed cold-induced sweetening resulting in minimum accumulation of reducing sugars in genome edited tubers.

The transcription factor StTINY3 enhances cold-induced sweetening resistance by coordinating starch resynthesis and sucrose hydrolysis in potato.

The accumulation of reducing sugars in cold-stored tubers, known as cold-induced sweetening (CIS), negatively affects potato processing quality. The starch to sugar interconversion pathways that are altered in cold-stored CIS tubers have been elucidated, but the mechanism that regulates them remains largely unknown. This study identified a CBF/DREB transcription factor (StTINY3) that enhances CIS resistance by both activating starch biosynthesis and repressing the hydrolysis of sucrose to reducing sugars in detached cold-stored tubers. Silencing StTINY3 in a CIS-resistant genotype decreased CIS resistance, while overexpressing StTINY3 in a CIS-sensitive genotype increased CIS resistance, and altering StTINY3 expression was associated with expression changes in starch resynthesis-related genes. We showed first that overexpressing StTINY3 inhibited sucrose hydrolysis by enhancing expression of the invertase inhibitor gene StInvInh2, and second that StTINY3 promoted starch resynthesis by up-regulating a large subunit of the ADP-glucose pyrophosphorylase gene StAGPaseL3, and the glucose-6-phosphate transporter gene StG6PT2. Using electrophoretic mobility shift assays, we revealed that StTINY3 is a nuclear-localized transcriptional activator that directly binds to the dehydration-responsive element/CRT cis-element in the promoters of StInvInh2 and StAGPaseL3. Taken together, these findings established that StTINY3 influences CIS resistance in cold-stored tubers by coordinately modulating the starch to sugar interconversion pathways and is a good target for improving potato processing quality.

Invertase inhibitors in potato: towards a biochemical and molecular understanding of cold-induced sweetening.

Invertase inhibitors classified as cell wall/apoplastic and vacuolar belonging to the pectin methylesterase family, play a major role in cold-induced sweetening (CIS) process of potato tubers. The CIS process is controlled at the post-translational level via an interaction between invertase (cell wall/apoplastic and vacuolar) by their compartment-specific inhibitors (cell wall/apoplastic and vacuolar). Invertase inhibitors have been cloned, sequenced and functionally characterized from potato cultivars differing in their CIS ability. The secondary structure of the invertase inhibitors consisted of seven alpha-helices and four conserved cysteine residues. The well-conserved three amino acids i.e. Pro-Lys-Phe are known to interact with invertase. Location of the genes encoding cell wall/apoplastic and vacuolar invertase inhibitors on potato chromosome number twelve in a tandem orientation without any intervening genes suggest their divergence into the cell wall and vacuole forms following the event of gene duplication. Under cold storage conditions, the vacuolar invertase inhibitor gene showed developmentally regulated alternative splicing and produce hybrid mRNAs which were the result of mRNA splicing of an upstream region of vacuolar invertase inhibitor gene to a downstream region of the apoplastic invertase inhibitor gene. Transgenic potato tubers overexpressing invertase inhibitors resulted in decreased invertase activity, low reducing sugars and improved processing quality making invertase inhibitors highly potential candidate genes for overcoming CIS. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene-editing technology offers transgene-free breeding for developing CIS resistant potato cultivars. Moreover, the post-transcriptional regulation of invertase inhibitors during cold storage can be warranted. This review summarizes progress and current knowledge on biochemical and molecular approaches used for the understanding of invertase inhibitors with special reference to key findings in potato.

Quantitative trait loci for starch-corrected chip color after harvest, cold storage and after reconditioning mapped in diploid potato.

The objective of this study was to map the quantitative trait loci (QTLs) for chip color after harvest (AH), cold storage (CS) and after reconditioning (RC) in diploid potato and compare them with QTLs for starch-corrected chip color. Chip color traits AH, CS, and RC significantly correlated with tuber starch content (TSC). To limit the effect of starch content, the chip color was corrected for TSC. The QTLs for chip color (AH, CS, and RC) and the starch-corrected chip color determined with the starch content after harvest (SCAH), after cold storage (SCCS) and after reconditioning (SCRC) were compared to assess the extent of the effect of starch and the location of genetic factors underlying this effect on chip color. We detected QTLs for the AH, CS, RC and starch-corrected traits on ten potato chromosomes, confirming the polygenic nature of the traits. The QTLs with the strongest effects were detected on chromosomes I (AH, 0 cM, 11.5% of variance explained), IV (CS, 43.9 cM, 12.7%) and I (RC, 49.7 cM, 14.1%). When starch correction was applied, the QTLs with the strongest effects were revealed on chromosomes VIII (SCAH, 39.3 cM, 10.8% of variance explained), XI (SCCS, 79.5 cM, 10.9%) and IV (SCRC, 43.9 cM, 10.8%). Applying the starch correction changed the landscape of QTLs for chip color, as some QTLs became statistically insignificant, shifted or were refined, and new QTLs were detected for SCAH. The QTLs on chromosomes I and IV were significant for all traits with and without starch correction.

Effects of (S)-Carvone and Gibberellin on Sugar Accumulation in Potatoes during Low Temperature Storage.

Potato tubers (Solanum tuberosum L.) are usually stored at low temperature, which can suppress sprouting and control the occurrence of diseases. However, low temperatures lead potatoes to easily suffer from cold-induced sweetening (CIS), which has a negative effect on food processing. The aim of this research was to investigate potential treatments on controlling CIS in potatoes during postharvest storage. "Atlantic" potatoes were treated with gibberellin and (S)-carvone, respectively, and stored at 4 °C for 90 days. The results showed that gibberellin can significantly accelerate sprouting and sugar accumulation by regulating expressions of ADP-glucose pyrophosphorylase (AGPase), granule-bound starch synthase (GBSS), ß-amylase (BAM1/2), UDP-glucose pyrophosphorylase (UGPase) and invertase inhibitor (INH1/2) genes. The opposite effects were found in the (S)-carvone treatment group, where CIS was inhibited by modulation of the expressions of GBSS and INH1/2 genes. In summary, gibberellin treatment can promote sugar accumulation while (S)-carvone treatment has some effects on alleviating sugar accumulation. Thus, (S)-carvone can be considered as a potential inhibitor of some of the sugars which are vital in controlling CIS in potatoes. However, the chemical concentration, treatment time, and also the treatment method needs to be optimized before industrial application.

Gene expression profiles predictive of cold-induced sweetening in potato.

Cold storage (2-4 °C) used in potato production to suppress diseases and sprouting during storage can result in cold-induced sweetening (CIS), where reducing sugars accumulate in tuber tissue leading to undesirable browning, production of bitter flavors, and increased levels of acrylamide with frying. Potato exhibits genetic and environmental variation in resistance to CIS. The current study profiles gene expression in post-harvest tubers before cold storage using transcriptome sequencing and identifies genes whose expression is predictive for CIS. A distance matrix for potato clones based on glucose levels after cold storage was constructed and compared to distance matrices constructed using RNA-seq gene expression data. Congruence between glucose and gene expression distance matrices was tested for each gene. Correlation between glucose and gene expression was also tested. Seventy-three genes were found that had significant p values in the congruence and correlation tests. Twelve genes from the list of 73 genes also had a high correlation between glucose and gene expression as measured by Nanostring nCounter. The gene annotations indicated functions in protein degradation, nematode resistance, auxin transport, and gibberellin response. These 12 genes were used to build models for prediction of CIS using multiple linear regression. Nine linear models were constructed that used different combinations of the 12 genes. An F-box protein, cellulose synthase, and a putative Lax auxin transporter gene were most frequently used. The findings of this study demonstrate the utility of gene expression profiles in predictive diagnostics for severity of CIS.

Cytosolic glyceraldehyde-3-phosphate dehydrogenases play crucial roles in controlling cold-induced sweetening and apical dominance of potato (Solanum tuberosum L.) tubers.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an important enzyme that functions in producing energy and supplying intermediates for cellular metabolism. Recent researches indicate that GAPDHs have multiple functions beside glycolysis. However, little information is available for functions of GAPDHs in potato. Here, we identified 4 putative cytosolic GAPDH genes in potato genome and demonstrated that the StGAPC1, StGAPC2, and StGAPC3, which are constitutively expressed in potato tissues and cold inducible in tubers, encode active cytosolic GAPDHs. Cosuppression of these 3 GAPC genes resulted in low tuber GAPDH activity, consequently the accumulation of reducing sugars in cold stored tubers by altering the tuber metabolite pool sizes favoring the sucrose pathway. Furthermore, GAPCs-silenced tubers exhibited a loss of apical dominance dependent on cell death of tuber apical bud meristem (TAB-meristem). It was also confirmed that StGAPC1, StGAPC2, and StGAPC3 interacted with the autophagy-related protein 3 (ATG3), implying that the occurrence of cell death in TAB-meristem could be induced by ATG3 associated events. Collectively, the present research evidences first that the GAPC genes play crucial roles in diverse physiological and developmental processes in potato tubers.

Amylases StAmy23, StBAM1 and StBAM9 regulate cold-induced sweetening of potato tubers in distinct ways.

Cold-induced sweetening (CIS) in potato is detrimental to the quality of processed products. Conversion of starch to reducing sugars (RS) by amylases is considered one of the main pathways in CIS but is not well studied. The amylase genes StAmy23, StBAM1, and StBAM9 were studied for their functions in potato CIS. StAmy23 is localized in the cytoplasm, whereas StBAM1 and StBAM9 are targeted to the plastid stroma and starch granules, respectively. Genetic transformation of these amylases in potatoes by RNA interference showed that ß-amylase activity could be decreased in cold-stored tubers by silencing of StBAM1 and collective silencing of StBAM1 and StBAM9. However, StBAM9 silencing did not decrease ß-amylase activity. Silencing StBAM1 and StBAM9 caused starch accumulation and lower RS, which was more evident in simultaneously silenced lines, suggesting functional redundancy. Soluble starch content increased in RNAi-StBAM1 lines but decreased in RNAi-StBAM9 lines, suggesting that StBAM1 may regulate CIS by hydrolysing soluble starch and StBAM9 by directly acting on starch granules. Moreover, StBAM9 interacted with StBAM1 on the starch granules. StAmy23 silencing resulted in higher phytoglycogen and lower RS accumulation in cold-stored tubers, implying that StAmy23 regulates CIS by degrading cytosolic phytoglycogen. Our findings suggest that StAmy23, StBAM1, and StBAM9 function in potato CIS with varying levels of impact.

UV-C treatment on physiological response of potato (Solanum tuberosum L.) during low temperature storage.

The storage of potato tuber (Solanum tuberosum L.) at low temperatures minimizes sprouting and disease but can cause cold-induced sweetening (CIS), which leads to the production of the cancerogenic substance acrylamide during the frying processing. The aim of this research was to investigate the effects of UV-C treatment on CIS in cold stored potato tuber. 'Atlantic' potatoes were treated with UV-C for an hour and then stored at 4 °C up to 28 days. The UV-C treatment significantly prevented the increase of malondialdehyde content (an indicator of the prevention of oxidative injury) in potato cells during storage. The accumulation of reducing sugars, particularly fructose and glucose, was significantly reduced by UV-C treatment possibly due to the regulation of the gene cascade, sucrose phosphate synthase, invertase inhibitor 1/3, and invertase 1 in potato tuber, which were observed to be differently expressed between treated and untreated potatoes during low temperature storage. In summary, UV-C treatment prevented the existence of oxidative injury in potato cells, thus, lowered the amount of reducing sugar accumulation during low temperature storage of potato tubers.

Improving cold storage and processing traits in potato through targeted gene knockout.

Cold storage of potato tubers is commonly used to reduce sprouting and extend postharvest shelf life. However, cold temperature stimulates the accumulation of reducing sugars in potato tubers. Upon high-temperature processing, these reducing sugars react with free amino acids, resulting in brown, bitter-tasting products and elevated levels of acrylamide--a potential carcinogen. To minimize the accumulation of reducing sugars, RNA interference (RNAi) technology was used to silence the vacuolar invertase gene (VInv), which encodes a protein that breaks down sucrose to glucose and fructose. Because RNAi often results in incomplete gene silencing and requires the plant to be transgenic, here we used transcription activator-like effector nucleases (TALENs) to knockout VInv within the commercial potato variety, Ranger Russet. We isolated 18 plants containing mutations in at least one VInv allele, and five of these plants had mutations in all VInv alleles. Tubers from full VInv-knockout plants had undetectable levels of reducing sugars, and processed chips contained reduced levels of acrylamide and were lightly coloured. Furthermore, seven of the 18 modified plant lines appeared to contain no TALEN DNA insertions in the potato genome. These results provide a framework for using TALENs to quickly improve traits in commercially relevant autotetraploid potato lines.

The potato amylase inhibitor gene SbAI regulates cold-induced sweetening in potato tubers by modulating amylase activity.

Potato cold-induced sweetening (CIS) is critical for the postharvest quality of potato tubers. Starch degradation is considered to be one of the key pathways in the CIS process. However, the functions of the genes that encode enzymes related to starch degradation in CIS and the activity regulation of these enzymes have received less attention. A potato amylase inhibitor gene known as SbAI was cloned from the wild potato species Solanum berthaultii. This genetic transformation confirmed that in contrast to the SbAI suppression in CIS-resistant potatoes, overexpressing SbAI in CIS-sensitive potatoes resulted in less amylase activity and a lower rate of starch degradation accompanied by a lower reducing sugar (RS) content in cold-stored tubers. This finding suggested that the SbAI gene may play crucial roles in potato CIS by modulating the amylase activity. Further investigations indicated that pairwise protein-protein interactions occurred between SbAI and α-amylase StAmy23, ß-amylases StBAM1 and StBAM9. SbAI could inhibit the activities of both α-amylase and ß-amylase in potato tubers primarily by repressing StAmy23 and StBAM1, respectively. These findings provide the first evidence that SbAI is a key regulator of the amylases that confer starch degradation and RS accumulation in cold-stored potato tubers.

Identification of potato varieties suitable for cold storage and reconditioning: A safer alternative to anti-sprouting chemicals for potato sprouting control.

Low temperature storage as an alternative to anti-sprouting chemicals in potato storage may induce reducing sugars (RS) accumulation (i.e. glucose and fructose) in potato tubers. This phenomenon is called "cold induced sweetening" (CIS) and occurs in certain varieties. CIS leads to a decrease in the organoleptic qualities and darkening of processed potato and the accumulation of toxic molecules such as acrylamide. To identify potato varieties suitable for storage at low temperatures, we screened six commercial processing varieties: Lady Claire (LC), Verdi, Kiebitz (KB), Pirol, Agria and Markies for their CIS characteristics and sprout-forming potential after storage at 4 °C and 8 °C. Our findings reveal that 4 °C storage allows for efficient sprout reduction in all six tested varieties for up to 4.5 months of storage. Three CIS-resistant varieties, namely Verdi, Lady Claire and Kiebitz, were identified as able to be stored for up to four months at 4 °C with limited increase in glucose content. Conversely, Pirol, Agria and Markies showed an increase in glucose content with a decrease in storage temperature and can be considered as CIS-susceptible varieties. After processing into crisps, the CIS-susceptible varieties displayed poor crisp color quality (brown to black color crisps) after storage for two months at 4 °C compared to the storage at 8 °C, whereas the CIS-resistant varieties had good crisp color quality (pale yellow color crisps) after storage at both 4 and 8 °C. Interestingly, the trends of total RS and/or glucose content in the CIS-resistant and in the CIS-susceptible varieties were correlated with the trends in Vacuolar Invertase (VInv) gene expression for most varieties, as well as with the trends in acrylamide content after processing. In addition, reconditioning of Markies variety after storage at 4 °C by gradually increasing the temperature to 15 °C resulted in a significant decrease of VInv transcript levels (reduction of 80 %), acrylamide content (reduction of 75 %) and glucose content when compared to a storage at 4 °C without reconditioning. Those results demonstrate that the reconditioning technique is a key factor for a sustainable potato storage and for improving the quality of processed potatoes.

Amelioration of cold-induced sweetening in potato by RNAi mediated silencing of StUGPase encoding UDP-glucose pyrophosphorylase.

Cold-induced sweetening (CIS) is an unwanted physiological phenomenon in which reducing sugars (RS) get accumulated in potato (Solanum tuberosum) upon cold storage. High RS content makes potato commercially unsuitable for processing due to the unacceptable brown color in processed products like chips, fries, etc., and the production of a potential carcinogen, acrylamide. UDP-glucose pyrophosphorylase (UGPase) catalyzes the synthesis of UDP-glucose towards the synthesis of sucrose and is also involved in the regulation of CIS in potato. The objective of the present work was RNAi-mediated downregulation of the StUGPase expression level in potato for the development of CIS tolerant potato. Hairpin RNA (hpRNA) gene construct was developed by placing UGPase cDNA fragment in sense and antisense orientation intervened by GBSS intron. Internodal stem explants (cv. Kufri Chipsona-4) were transformed with hpRNA gene construct, and 22 transgenic lines were obtained by PCR screening of putative transformants. Four transgenic lines showed the highest level of RS content reduction following 30 days of cold storage, with reductions in sucrose and RS (glucose & fructose) levels of up to 46% and 57.5%, respectively. Cold stored transgenic potato of these four lines produced acceptable chip colour upon processing. The selected transgenic lines carried two to five copies of the transgene. Northern hybridization revealed an accumulation of siRNA with a concomitant decrease in the StUGPase transcript level in these selected transgenic lines. The present work demonstrates the efficacy of StUGPase silencing in controlling CIS in potato, and the strategy can be employed for the development of CIS tolerant potato varieties.

Comparative transcriptome reveals distinct starch-sugar interconversion patterns in potato genotypes contrasting for cold-induced sweetening capacity.

Potato accumulates large amounts of soluble sugar during cold storage periods. However, a system based understanding of this process is still largely unknown. Here, we compared the dynamic cold-responded transcriptome of genotypes between cold-induced sweetening resistant (CIS-R) and cold-induced sweetening sensitive (CIS-S) in tubers. Comparative transcriptome revealed that activating the pathways of starch degradation, sucrose synthesis and hydrolysis could be common strategies in response to cold in both genotypes. Moreover, the variation in sugar accumulation between genotypes may be due to genetic differences in cold response, which could be mainly explained: CIS-R genotype was active in starch synthesis and attenuated in sucrose hydrolysis by promoting the coordinate expression of aseries ofgenes involved in starch-sugar interconversion. Additionally, transcription factors, the candidate master regulators of starch-sugar interconversion, were discussed. Taken together, this work has provided an avenue for studying the mechanism involved in the regulation of the CIS resistance.

Prospects for potato genome editing to engineer resistance against viruses and cold-induced sweetening.

Crop improvement through transgenic technologies is commonly tagged with GMO (genetically-modified-organisms) where the presence of transgene becomes a big question for the society and the legislation authorities. However, new plant breeding techniques like CRISPR/Cas9 system [clustered regularly interspaced palindromic repeats (CRISPR)-associated 9] can overcome these limitations through transgene-free products. Potato (Solanum tuberosum L.) being a major food crop has the potential to feed the rising world population. Unfortunately, the cultivated potato suffers considerable production losses due to several pre- and post-harvest stresses such as plant viruses (majorly RNA viruses) and cold-induced sweetening (CIS; the conversion of sucrose to glucose and fructose inside cell vacuole). A number of strategies, ranging from crop breeding to genetic engineering, have been employed so far in potato for trait improvement. Recently, new breeding techniques have been utilized to knock-out potato genes/factors like eukaryotic translation initiation factors [elF4E and isoform elF(iso)4E)], that interact with viruses to assist viral infection, and vacuolar invertase, a core enzyme in CIS. In this context, CRISPR technology is predicted to reduce the cost of potato production and is likely to pass through the regulatory process being marker and transgene-free. The current review summarizes the potential application of the CRISPR/Cas9 system for traits improvement in potato. Moreover, the prospects for engineering resistance against potato fungal pathogens and current limitations/challenges are discussed.

Tuber transcriptome profiling of eight potato cultivars with different cold-induced sweetening responses to cold storage.

Tubers are vegetative reproduction organs formed from underground extensions of the plant stem. Potato tubers are harvested and stored for months. Storage under cold temperatures of 2-4 °C is advantageous for supressing sprouting and diseases. However, development of reducing sugars can occur with cold storage through a process called cold-induced sweetening (CIS). CIS is undesirable as it leads to darkened color with fry processing. The purpose of the current study was to find differences in biological responses in eight cultivars with variation in CIS resistance. Transcriptome sequencing was done on tubers before and after cold storage and three approaches were taken for gene expression analysis: 1. Gene expression correlated with end-point glucose after cold storage, 2. Gene expression correlated with increased glucose after cold storage (after-before), and 3. Differential gene expression before and after cold storage. Cultivars with high CIS resistance (low glucose after cold) were found to increase expression of an invertase inhibitor gene and genes involved in DNA replication and repair after cold storage. The cultivars with low CIS resistance (high glucose after cold) showed increased expression of genes involved in abiotic stress response, gene expression, protein turnover and the mitochondria. There was a small number of genes with similar expression patterns for all cultivars including genes involved in cell wall strengthening and phospholipases. It is proposed that the pattern of gene expression is related to chilling-induced DNA damage repair and cold acclimation and that genetic variation in these processes are related to CIS.