Project description:Dormant/sprouting bud or eye tissue was collected from field grown Russet Burbank tubers using melon baler. These tubers after harvest washed with 5% Clorox, dried and stored at room temperature and allowed to sprout. Tissue samples were collected from dormant and sprouting eye at different physiological stages (based on length of the sprout) of sprouting. RNA was extracted using a hot phenol method and treated with DNAse. RNA extracted from different stages of sprouting potato tubers was used as query samples. RNA collected from non-sprouting eyes of potato tubers was used as reference samples Information on RNA samples. Plant species: S. tuberosum CV Russet Burbank. Tissue harvested: Tissue was harvested from the dormant/sprouting bud/eye using ½ inch melon baler. Time points: Dormant eyes –Stage 1 Initiating buds/sprouts – Stage 2 Sprouts (1/8 inch) – Stage 3 Sprouts (1/4 inch) – Stage 4 Sprout callus (1/4 inch) – Stage 5 Sprouts (1/2 inch) – Stage 6. Growth conditions: Field grown. Replicate information: Biological replicates; A, B and C Keywords: Direct comparison
Project description:An in vivo and in vitro potato tuber development gene expression study. For in vitro tuber development expression analysis, RNA was isolated from in vitro microtubers at 2, 5, 10, 20 and 30 days following observed tuber induction. Two microtuber populations were used as biological replicates for the developmental stages. The RNA from all developmental stages was pooled to generate the reference samples. Ten microarray hybridizations were performed. For in vivo tuber development expression analysis, RNA was isolated from tubers growing in growth chamber conditions. Tissues were divided into six group, according to developmental size: stolon (no tuber formation), 1-5 mm tubers, 6-10 mm tubers, 11-15 mm tubers, 16-25 mm tubers, and 26-35 mm tubers. Two biological replicates of ten plants each were grown sequentially in the same growth chamber. The RNA from all developmental stages was pooled to generate the reference samples. Twelve microarray hybridizations were performed. For all experiments, the RNA was labeled using the indirect labeling method with random hexamer primers. Amplified cRNA was used as labeling template for stolons. Total RNA was used as labeling template in all other labeling reactions.
Project description:Blackspot bruising is the tuber discoloration resulting from mechanical damage in susceptible crops and is a major quality and environmental problem for the potato industry. The present study is aimed at identifying differences in gene expression which are correlated with the development of bruise susceptibility or resistance in tubers. This may produce markers or diagnostic tests for use in predicting enhanced bruise susceptibility in field grown potato crops. Different potato cultivars were grown in field plots at ADAS Gleadthorpe, Nottinghamshire, UK during three growing seasons – 2003, 2004 and 2005. Random samples of tubers (8) were harvested by hand and cores excised from the subdermal tissues of the stolon end of the tubers and immediately frozen and stored in liquid nitrogen for RNA isolations. The remainder of the crops were stored at 4oC for bruise susceptibility testing - samples of tubers (30) from each crop were impacted and incubated for 48h at 30oC. Bruise susceptibility was scored, according to the intensity of pigmentation and the volume of tuber tissue affected, on a scale of 0 (very resistant) – 10 (very susceptible). Crops representing 'very susceptible', 'susceptible', 'moderate' and 'resistant' to bruising, were selected for RNA isolation. Total RNA’s were isolated from frozen cores by the TIGR hot phenol extraction method, precipitated by LiCl and treated with DNase. RNA concentrations were adjusted to ~2ug/ul and analysed on 1% TBE gels after formamide treatment. Any RNA’s which were not pure enough or showed degradation were discarded. Total RNA’s were pooled and 25ul samples aliquotted according to the required hybridisations indicated in the Excel spreadsheet and the on-line information form. Keywords: Direct comparison
Project description:Blackspot bruising is the tuber discoloration resulting from mechanical damage in susceptible crops and is a major quality and environmental problem for the potato industry. The present study is aimed at identifying differences in gene expression which are correlated with the development of bruise susceptibility or resistance in tubers. This may produce markers or diagnostic tests for use in predicting enhanced bruise susceptibility in field grown potato crops. Different potato cultivars were grown in field plots at ADAS Gleadthorpe, Nottinghamshire, UK during three growing seasons – 2003, 2004 and 2005. Random samples of tubers (8) were harvested by hand and cores excised from the subdermal tissues of the stolon end of the tubers and immediately frozen and stored in liquid nitrogen for RNA isolations. The remainder of the crops were stored at 4oC for bruise susceptibility testing - samples of tubers (30) from each crop were impacted and incubated for 48h at 30oC. Bruise susceptibility was scored, according to the intensity of pigmentation and the volume of tuber tissue affected, on a scale of 0 (very resistant) – 10 (very susceptible). Crops representing 'very susceptible', 'susceptible', 'moderate' and 'resistant' to bruising, were selected for RNA isolation. Total RNA’s were isolated from frozen cores by the TIGR hot phenol extraction method, precipitated by LiCl and treated with DNase. RNA concentrations were adjusted to ~2ug/ul and analysed on 1% TBE gels after formamide treatment. Any RNA’s which were not pure enough or showed degradation were discarded. Total RNA’s were pooled and 25ul samples aliquotted according to the required hybridisations indicated in the Excel spreadsheet and the on-line information form. Keywords: Direct comparison 32 hybs total
Project description:The goal of the current research is to identify factors that involved with heat induced russeting of the potato tuber skin. Potato plants of the variety Desirèe were grown in pots filled with perlite, in a greenhouse under natural winter conditions (Nov 2005- Jan 2006, average temperatures range of 10-18°C). For the exposure of tubers to heat stress (H) hot water (33-35°C) was circulated in tubes lined at the internal side of the pots. The heat was applied one week before tubers harvest. Tubers were harvested at two time points: 8 weeks post sprouting and a week post mechanical vine killing. The skin (S) of young tubers was peeled by hand, as the remaining phelloderm (PH) layers, the periderm of young tubers (P) and the periderm of mature tubers (ST) were peeled using a scalpel blade. Leaves (L) and tuber flesh (TF) samples were collected as well. For each RNA sample 4 biological replicates were prepared; each one represents pooled tissues from 4-6 plants grown at different location in the greenhouse. RNA was extracted using CTAB protocol, and was further purified by RNeasy Mini Kit (Qiagen) using the On-Column DNase Digestion protocol. Keywords: Loop design
Project description:Anthocyanins are plant pigments responsible for the colors of many flowers, fruits and storage organs and have roles in abiotic and biotic stress resistance. Anthocyanins and polyphenols are bioactive compounds in plants including potato (Solanum tuberosum L.) which is the most important non-cereal crop in the world, cultivated for its tubers rich in starch and nutrients. The genetic regulation of the flavonoid biosynthetic pathway is relatively well known leading to the formation of anthocyanins. However, our knowledge of post-transcriptional regulation of anthocyanin biosynthesis is limited. There is increasing evidence that micro RNAs (miRNAs) and other small RNAs can regulate the expression level of key factors in anthocyanin production. In this study we have found strong associations between the high levels of miR828, TAS4 D4(-) and purple/red color of tuber skin and flesh. This was confirmed not only in different cultivars but in pigmented and non-pigmented sectors of the same tuber. Phytochemical analyses verified the levels of anthocyanins and polyphenols in different tissues. We showed that miR828 is able to direct cleavage of the RNA originating from Trans-acting siRNA gene 4 (TAS4) and initiate the production of phased small interfering RNAs (siRNAs) whose production depends on RNA-dependent RNA polymerase 6 (RDR6). MYB transcription factors were predicted as potential targets of miR828 and TAS4 D4(-) and their expression was characterized. MYB12 and R2R3-MYB genes showed decreased expression levels in purple skin and flesh in contrast with high levels of small RNAs in the same tissues. Moreover, we confirmed that R2R3-MYB and MYB-36284 are direct targets of the small RNAs. Overall, this study sheds light on the small RNA directed anthocyanin regulation in potato, which is an important member of the Solanaceae family.
Project description:Potato (Solanum tuberosum L) is a natural host of Potato spindle tuber viroid (PSTVd) which can cause characteristic symptoms on developing plants including stunting phenotype and distortion of leaves and tubers. PSTVd is the type species of the family Pospiviroidae, it can replicate in the nucleus and the viroid RNA moves systemically in infected plants. Its KF440-2 strain can cause severe symptoms in potato. It is not well understood how the viroid can affect host genes for successful invasion and which genes show altered expression levels upon infection. In this study, we used a high-scale method to identify differentially expressed genes in potato. We have identified defence, stress and sugar metabolism related genes having altered expression levels upon infection. Additionally, hormone pathways connected genes showed up- or down-regulation. Our primary focus is on the identification of genes which can affect tuber formation as the viroid infection can strongly influence tuber development, especially tuber shape is affected. DWARF1/DIMINUTO, Gibberellin 7-oxidase and BEL5 protein were identified and validated which showed differential expression in viroid infected tissues suggesting that gibberellin and brassinosteroid pathways have a possible role in tuber development upon PSTVd infection.
Project description:Effects of different parameters on the transcriptome in potato tuber: effect of infection with potato virus Y (PVY) on potato tubers, effects of two different storage times of potato tubers compared to no storage, effect of different storage temperature on potato tubers, effect of tuber necrosis development, effects of interactions between the above parameters. Lists of interaction factors and the differentially-expressed genes associated with each factor are provided as a series of Additional Files to this submission (see http://www.ebi.ac.uk/arrayexpress/files/E-MTAB-1071).
Project description:To extend our understanding of systemic necrosis in susceptible potato tubers infected with the necrotic strain of Potato virus Y (PVYNTN) gene expression was compared between healthy and infected non-necrotic and necrotic (both non-necrotic and necrotic tissue) potato tubers.