Illumina Genome Analyzer sequencing; Transcriptome Analysis of the potato (genotype RH89-039-16)
Transcriptome Analysis of the potato (genotype RH89-039-16)
RH tubers were planted in soil-filled pots in the greenhouse. After plant emergence and several weeks of growth, non-tuberizing stolons were harvested (RH Stolon). Around one week after the first swellings were observed, small tubers were collected with a maximum size of 1cm (RH Young tuber). Around the same time, anthesis occurred and both open flowers and developing flower buds were harvested (RH flower). Around the 12th leaf stage (4 weeks after first visible swelling) various tissues were harvested, including leaves (fully expanded), shoot apex, petioles, stem, tubers (RH Mature Tuber) and roots, and immediately frozen in liquid nitrogen. Stamens were collected at the same time from fully opened flowers. All tissue samples were collected from at least five different plants. For the water-stressed leaf tissue sample, a subset of plants was denied water for 2 days after which wilting leaves were harvested (2nd - 4th fully expanded leaf). For the tuber cross section, plants were left to senescence. Tubers were then collected, and peel, cortex and pith tissues were sampled and immediately frozen in liquid nitrogen. Sprouts of previously harvested RH tubers were harvested after storage at RT in the dark for 3-4 months. RH in vitro plants were grown on standard MS medium on 2% sucrose with 16 h of light at 24C for 2 weeks, after which the whole explant, including roots, was harvested. A total of 16 tissues was collected and RNA was isolated using the RNeasy kit (Qiagen), except for the tuber and root tissues, which were isolated using the hot-phenol method. The latter samples were than purified using the RNAeasy columns. For all samples, on-column DNaseI treatment was performed and the quality and integrity of the obtained RNA was checked using the ND-100 spectrophotometer (Nanodrop Technologies) and agarose gel-electrophoresis.
Experimental Factor: ORGANISMPART
flower