GSM1174035: WT6; Homo sapiens; RNA-Seq
The captured leukocytes were treated with RNAlater immediately after filtration of the blood and were maintained frozen until RNA extraction. RNA quality was assessed by running the samples on Agilent RNA 6000 Nano gels (#5067-1511). For each library, ribosomal RNA of 5 ug total RNA was removed using the Invitrogen RiboMinus kit (# A10837-08), and then the sample was concentrated using the RiboMinus™ Concentration Module (Invitrogen). Ribosomal RNA removal was verified by RNA 6000 Nano gel analysis. Library construction was done according to the SOLiD™ Whole Transcriptome Analysis Kit (PN 4425680) protocol. Fragmentation (by RNase III) was verified on Agilent RNA 6000 Pico Kit (#5067-1513), and 150 ng fragmented RNA were used for further protocol. cDNA samples were run on 4% agarose gel, 150-250bp fragments were cut and extracted using the Qiagen Min-Elute Gel Extraction Kit (#28604), and the gel was dissolved by intensive vortexing and not by heating. Libraries were barcoded and amplified for 12 cycles using barcoded primers supplied in the SOLiD™ Transcriptome Multiplexing Kit (Ambion 4427046). Libraries were quantified using the Kapa ABI SOLiD Library Quantification Kit (KK4833) and diluted for final analysis on Agilent High Sensitivity DNA Kit (#5067-4626). 500 uM libraries were used for emulsion PCR according to the Applied Biosystems SOLiD™ 3 System Templated Bead Preparation Guide (4407421 ) to prepare for sequencing on the SOLiD 3 platform.
GEO Accession
GSM1174035