GSM2887490: CTS_175; Homo sapiens; RNA-Seq
Specimens were immersed in RNAlaterTM Stabilization Solution (Thermo Fisher Scientific) and stored at -80°C. RNA was extracted using a combination of the phenol extraction and column purification. Briefly, samples were homogenised in Trizol (Invitrogen), mixed with chloroform and centrifuged in order to collect the aqueous liquid phase. This solution was placed on High Pure RNA Isolation Kit (Roche) columns and subjected to multiple washes to purify the RNA sample, as per the manufacturers instructions. Library preparation was polyA enriched and directional, using Illumina TruSeq Stranded mRNA and standard universal Illumina multiplexing adapters: 5' P-GATCGGAAGAGCACACGTCT and 5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT. The poly-A selected RNA was converted to cDNA using the strand-specific (deoxy-UTP strand-marking protocol) dUTP (Borodina et al., 2011) protocol. Amplification was performed by unique dual indexing using indexing primers described elsewhere71. Sequencing was performed using HiSeq4000 with 75bp read length and paired end reads. Healthy skin samples and CTS tenosynovium samples were extracted and sequenced using the same protocol but in different library batches.    Oxford Genomics produced FastQ sequencing files which encode quality metrics following the Sanger standard, i.e. Sanger qualities, using the standard Phred score (Ewing and Green, 1998). All sequencing lanes gave high yield, consistent GC content, consistent and expected sequence insert between the paired-end adapters and high quality base calling.  Individuals were multiplexed in lanes.
GEO Accession
GSM2887490