GSM8446964: elt-1_RNAi, smRNA-seq,bio rep1; Caenorhabditis elegans; ncRNA-Seq
Small RNAs were purified by mir-Vana miRNA isolation kit (Thermo Scientific, AM1560) from extracted Total RNAs of prepared worms. Small-RNA cloning was performed. Briefly, RNA 5' Pyrophosphohydrolase (RppH) (NEB, M0356S) was used for trimming 5' triphosphate of endogenous 22G-RNAs to monophosphate. The 3' and 5' adaptors were ligated sequentially using truncated ligase 2 (NEB, M0373S) and ligase 1 (NEB, M0437M). cDNA was then generated by Superscript III reverse transcriptase (Invitrogen,18064022) and amplified by Q5 polymerase (NEB, M0491S) with adapter sequences added at this step. Amplified PCR products were subjected to 12.5% native PAGE and size selection. Finally, High throughput sequencing was carried out using multiple Illumina platforms.