title: Healthy1
design: Nucleic acid isolation was carried out from 5ml of serum at Genotypic Technology s Genomics facility (www.genotypic.co.in) using Qiamp Circulating Nucleic acid kit according to the manufacturer s instructions (Bangalore, India). The isolated RNA was analyzed using a bioanalyzer (Agilent, CA, USA) to check for their quantity and integrity. Small RNA libraries for sequencing were prepared using the Illumina TruSeq Small RNA library kit according to the manufacturer s instructions. Briefly, 3  adaptors were ligated to the specific 3 OH group of RNAs 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, barcoded by PCR (21 cycles) and cleaned. The library was size selected in the range of 140   160 bp (corresponding to cDNAs from miRNAs) followed by overnight gel elution and salt precipitation using glycogen, 3M Sodium Acetate and absolute ethanol. The precipitate was re-suspended in nuclease free water. The prepared library was quantified (Qubit Thermo Fischer Scientific, USA) and validated for quality by running on a High Sensitivity Bioanalyzer Chip (Agilent, CA, USA). The libraries were sequenced using the Illumina HiSeq platform (Next Seq500) (CA, USA).