RNA-Seq for human brain: medial prefrontal cortex(MFC): Sample HSB149_MFC_L_mRNA
Tissue was pulverized with liquid nitrogen in a chilled mortar and pestle and transferred to a chilled safe-lock microcentrifuge tube. Per tissue mass, equal mass of chilled stainless steel beads along with two volumes of lysis buffer were added. Tissue was homogenized for 1 min in Bullet Blender and incubated at 37°C for 5 min. Lysis buffer up to 0.6 ml was again added, tissue homogenized for 1 min and incubated at 37°C for 1 min. Extraction was further carried out according to manufacturer’s protocol. Genomic DNA was removed by a proprietary column provided in RNeasy Plus Mini Kit (Qiagen) or by DNase treatment using TURBO DNA-free Kit. The mRNA-sequencing Sample preparation Kit (Illumina) was used to prepare cDNA libraries per manufacturer instructions.  The mixture of mRNA and spike-in RNAs was subjected to fragmentation, reverse transcription, end repair, 3’ end adenylation, and adapter ligation to generate libraries of short cDNA molecules, followed by PCR amplification.   mRNA-sequencing (mRNA-seq) was carried out on Illumina Genome Analyzer IIx (GAIIx),  according to the manufacturer's instructions.
alignment_software
STAR 2.4.0e