title: GSM3325605: MDA-MB-231 breast cancer cells coculture with 3T3-L1 adipocytes; Homo sapiens; miRNA-Seq
library_construction_protocol: Total RNA inclduing small RNA was extracted from MDA-MB-231 cells and subjected to small RNA deep sequencing Small RNA sequencing libraries were prepared with TruSeq Small RNA Sample Preparation Guide at the Genotypic Technology Pvt. Ltd., Bangalore, India. Briefly, approx. 50 ng of RNA was used as starting material. 3' adaptors were ligated to the specific 3'OH group of microRNAs followed by 5' adaptor ligation. The ligated products were reverse transcribed with superscript III reverse transcriptase by priming with reverse transcriptase primers. The cDNA was enriched and barcoded by PCR (15 cycles) and cleaned using polyacrylamide gel. The library was size selected in the range of 140 bp– 160 bp followed by overnight gel elution and salt precipitation using glycogen, 3 M sodium acetate and absolute ethanol.The precipitate was re-suspended in nuclease free water. The Illumina-compatible sequencing library was initially quantified by qubit fluorometer and its fragment size distribution was analyzed on Agilent Bioanalyzer. Finally, the sequencing library was accurately quantified by quantitative PCR using Kapa Library Quantification Kit. The qPCR quantified library was subjected to sequencing on an Illumina NextSeq 500 sequencer for 75 bp single-end chemistry.
attributes.tag: GEO Accession
attributes.value: GSM3325605