概要: To identificate long noncoding RNAs in rice, we profiled transcriptome of various organs at different developmental stages using stranded single-end RNA-seq based on poly(A) selection.
项目整体设计: Transcriptom profiling in flower buds, flowers, flag leaves and roots sampled before flowering and after flowering, milk grains and mature seeds.
Seeds from the cultivated rice subspecies Oryza sativa L. ssp. Japonica were grown in a greenhouse in Singapore . Flower buds were collected before flowering and flowers were collected at the flowering day. Flag leaves and roots were collected at the before- and after-flowering stage. The before-flowering sample was defined as a mixture of different stages in a period from panicle initiation to 1 day before flowering. The after-flowering sample was defined as a mixture of different stages after the flowering day. Milk grains and mature seeds were also collected.
处理方案:
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提取方案:
RNA was extracted using the Qiagen RNeasy Plant Mini kit. Total RNA was treated with Turbo DNase I (Life Technologies AM2238) according to product specification. DNase I treated RNA was then applied again to an RNeasy spin column with 0.5 volumes of ethanol, washed and eluted according to the manufacturer’s instructions.
建库方案:
Sequencing libraries were prepared using the Illumina Stranded mRNA sample Prep kit, set A (RS-122-2101, Illumina Inc.) according to the manufacturer's instructions. The quality and size of cDNA libraries for sequencing were checked using the Agilent 2200 TapeStation system (Agilent Inc.). The libraries were sequenced for 100 cycles on HiseqTM 2500 (Illumina Inc.)
