title: GSM1977381: WT_III; Ustilago maydis; RNA-Seq
library_construction_protocol: Samples were ground to a fine powder in liquid nitrogen and total RNAs were extracted using the TRIzolâ method (Invitrogen) Sequencing libraries were prepared with the Illumina TruSeq® RNA sample preparation Kit starting from 2µg total RNA. After poly-A selection (using poly-T oligo-attached magnetic beads), mRNA was purified and fragmented using divalent cations under elevated temperature. The RNA fragments were reverse transcribed using random primers. A second strand cDNA synthesis was carried out with DNA Polymerase I and RNase H. After end repair and A-tailing, indexing adapters were ligated to the cDNA. The products were then purified and amplified (14 PCR cycles) to create the final cDNA libraries. After library validation and quantification (Agilent 2100 Bioanalyzer), equimolar amounts of library were pooled. The pool was quantified using the Peqlab KAPA Library Quantification Kit and the Applied Biosystems 7900HT Sequence Detection System. The pool was sequenced using an Illumina TruSeq PE Cluster Kit v3 and an Illumina TruSeq SBS Kit v3-HS on an Illumina HiSeq 2000 sequencer with a paired-end (101x7x101 cycles) protocol.
attributes.tag: GEO Accession
attributes.value: GSM1977381