GSM5220545: miRNA isolated from non-treated sample; Arabidopsis thaliana; miRNA-Seq
Total RNA was isolated from the frozen calli of A. thaliana using TRIzol reagent Libraries were constructed following the High-Throughput illumina sequencing library protocol. After being checked for integrity, 40µg of total RNA was used to purify the poly-A-containing mRNA molecules using poly-T oligo-attached magnetic beads. Following purification, the mRNA is fragmented into small pieces using divalent cations under elevated temperature. Small RNA are selected by PAGE gel (separate 18-30nt RNA segment). The cleaved RNA fragments are used to synthesize cDNA strands and to amplify them using using a high-ping polymerase and enriched with both 3' and 5' adapter.  Library fragments (PCR products of 100~120 bp) were selected by PAGE to eliminate primer, dimer and other byproducts. Separation of the obtained fragments and the cyclization of single-strand DNA have occurred under high temperature. ssDNA circle was used to develop DNA nanoballs (DNBs).