GSM1842539: plasma_cancer_9; Homo sapiens; ncRNA-Seq
50 µl serum was mixed well with 12.6 µl ExoQuick exosome isolation solution, and incubated on ice for 30 min. Then the samples were centrifuged at 1000 g for 30 min at 4°C. The pellet was washed wish 1X DPBS and resuspended in 50 µl 1X DPBS. 20 µl samples were loaded onto the nanochip and incubated at 37ºC for 2 h. On each chip, two wells with 1XDPBS were used as negative control. The NextGen sequencing libraries were prepared using NEBNext Multiplex Small RNA Library Prep Set for Illumina sequencing kit (New England Biolabs). The 3’ and 5’ adapters and the reverse transcription primer were diluted in nuclease-free water to concentrations specified by the kit.  The adapter sequences were ligated to the 3’ and 5’ ends of the RNA isolated from 100 μL of plasma using T4 RNA ligase. Ligation reactions were performed overnight at 16oC to maximize efficiency. cDNA was then transcribed from the ligation product followed by PCR amplification with a unique barcode sequence for each sample. The fragment size of about 150 nts, was then extracted and purified from an acrylamide gel. The final purified and barcoded libraries were then pooled in equal molar for single-end 50 nt sequencing run using an Illumina HiSeq2500 instrument.
GEO Accession
GSM1842539