GSM6622362: microRNA, PASI >10, severe2; Homo sapiens; miRNA-Seq
Namely, smallRNA molecules were isolated from 1μg of total RNA via the adapter ligation. The isolated smallRNAs were synthesized as single-stranded cDNAs through the RT (Reverse transcription) priming. By applying this as a template for second strand synthesis, double-stranded cDNA was prepared through PCR (Polymerase Chain Reaction). And, the fragments around 150bp were extracted for sequencing through size selection by gel electrophoresis. Quality of these cDNA libraries was evaluated with the Agilent 2100 BioAnalyzer (Agilent, CA, USA) followed by quantification with the KAPA library quantification kit (Kapa Biosystems, MA, USA) according to the manufacturer's protocol. Following cluster amplification of denatured templates, sequencing was progressed using Illumina sequencing NovaSeq6000 S4 platform (Illumina, CA, USA). The libraries were prepared for 50bp single-end sequencing using the NEXTflex Small RNA-Seq Kit v3 (Bioo Scientific Corp.).