GSM2417587: total small RNAs in susi-1(R457H); Caenorhabditis elegans; miRNA-Seq
Total small RNAs were isolated from bleached embryos. Briefly, embryos were harvested and dounced on ice with pestle B in TRIzol reagent (Invitrogen) followed by isopropanol precipitation. The precipitates were treated with calf intestinal alkaline phosphatase (CIAP, Invitrogen), re-extracted with TRIzol, and treated with T4 polynucleotide kinase (T4 PNK, New England Biolabs) in the presence of 1 mM ATP. Small RNAs were cloned and deep sequenced using an Illumina platform, according to the manufacturer's instructions, by the Beijing Genomics Institute (BGI Shenzhen). sRNAs ranging from 20 to 30 nt were gel-purified and ligated to a 3’ adaptor (5’-pUCGUAUGCCGUCUUCUGCUUGidT-3’; p, phosphate; idT, inverted deoxythymidine) and a 5’ adaptor (5’-GUUCAGAGUUCUACAGUCCGACGAUC-3’). The ligation products were gel-purified, reverse transcribed, and amplified using Illumina’s sRNA primer set (5’-CAAGCAGAAGACG GCATACGA-3’; 5’-AATGATACGGCGACCACCGA-3’). The samples were sequenced using an Illumina Hiseq platform.
GEO Accession
GSM2417587