GSM2200238: Col_Al6h; Arabidopsis thaliana; RNA-Seq
Total RNA was isolated using the RNeasy® Plant Mini Kit (QIAGEN GmbH, Qiagen, GmbH, Hilden, Germany) following the manufacturer’s protocol.  RNA libraries were prepared for sequencing using standard Illumina protocols The total RNA samples are first treated with DNase I to degrade any possible DNA contamination. Then the mRNA is enriched by using the oligo(dT) magnetic beads (for eukaryotes). Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments (about 200 bp). Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3'-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to qualify and quantify of the sample library. The library products are ready for sequencing via Illumina HiSeqTM 2000 or other sequencer when necessary.
GEO
GSM2200238