GSM2342731: 59S2; Equus caballus; miRNA-Seq
Total RNA was extracted using Direct-zol RNA MiniPrep Kit (Zymo Research). The obtained RNA was then additionally purified and concentrated with RNA Clean & Concentrator™-5 (Zymo Research). 1ug of total RNA was used to prepare libraries. Libraries were prepared according to TruSeq Small RNA Sample Preparation Kit protocol (Illumina, protocol 15004197 Rev.F, February 2014). First, the samples were ligated with 3' and 5' adapters, respectively. Then, miRNA-adapter ligation products were reverse transcribed and PCR amplified (14 cycles). The amplification was carried out with the use of the universal primer and one of 12 different indexed primers for each sample. Then, the libraries' fragments of ~150bp (miRNA+adapters+PCR primer sequences) were band isolated from Novex 6% TBE PAGE gel (Invitrogen). The purified libraries were then captured and clustered on an Illumina flow cell. Finally, the obtained clustered libraries were sequenced on HiScanSQ platform (Illumina) following the manufacturer's protocols.
GEO Accession
GSM2342731