Escherichia coli K-12 small RNA sequencing
The strand-specific library was prepared and sequenced by Fasteris SA (Geneva, Switzerland) according to the Illumina TruSeq Small RNA Sample Prep protocol (2013). Shortly, RNA were extracted at the onset of stationary phase from cells grown in M9 minimal medium plus 0.2% glucose at 28C using the hot-phenol extraction procedure. 3g of RNA sample were loaded on a denaturing polyacrylamide gel (Mini PROTEAN 10% TBE gel, Bio-rad) after 5 min of denaturation at 65C and cooling on ice. After separation by electrophoresis, RNA of size ranging from 200 and 50 nucleotides were selected, extracted from the gel and precipitated. Then, the standard Illumina protocol was performed to prepare the strand-specific library of small RNA using the Illumina index n32. The cDNA was amplified by 10 duplication cycles of PCR, the size of the library was checked by electrophoresis and gel-purified to remove pairs of adaptors.
SUB4504766
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