GSM1973843: MNxMN_seedcoat1; Arabidopsis lyrata; RNA-Seq
Integenex PolyA prep protocol (Wafergen Biosystems) sequenced on 8/8/14 with barcode CTTGTA Seeds from crosses were hand-dissected into embryo, endosperm and seed coat, and total RNA was isolated using the RNAqueous Micro Kit (Ambion, Life Technologies Corp.). Input for mRNA-seq library construction varied from 120ng - 800ng. For bisulfite-sequencing libraries, genomic DNA was isolated from fresh flower bud tissue using the DNeasy Plant Mini Kit (Qiagen), or from dissected embryo or endosperm using CTAB. DNA was RNase treated. Strand-specific mRNA-seq libraries were prepared by the Whitehead Institute Genome Technology Core using the Integenex PolyA prep protocol (Wafergen Biosystems, Fremont CA) or using the Illumina TruSeq Stranded mRNA Kit. Libraries were multiplexed and sequenced on an Illumina HiSeq 2000 machine as 40x40, 80x80 or 100x100 paired end reads. 500 ng of genomic DNA was used for bisulfite treatment with the MethylCode Bisulfite Conversion Kit (Invitrogen, Life Technologies Corporation), according to the manufacturer’s protocol. Bisulfite-sequencing libraries were constructed using the EpiGnome Methyl-seq Kit (Epicentre, Biotechnologies, Madison, WI) according to the manufacturer’s protocol. Library quality was evaluated using a bioanalyzer and then sequenced on an Illumina HiSeq2000 using a 40x40 or 100x100 paired end protocol at the Whitehead Institute Genome Technology Core. Barcodes and date sequenced are indicated above.