sRNA sequencing of Lactobacillus casei BL23

Total RNR was extracted using RNAzol RT (RN190) (Molecular Research Center,Inc.) from L. casei BL23 grown at 37 C in BD (TM) Difc (TM) Lactobacilli MRS Broth collected at six different growth points and treated with DNaseI, RNase free (Thermo Scientific). rRNAs were depleted using modified RiboMinus Transcriptome Isolation Kit, bacteria (Invitrogen) and 50-500 nt RNA fraction was retrieved after size selection using 8 % denaturing PAA gel. After treatment with RNA 5' Pyrophosphohydrolase (RppH) (New England Biolabs) three biological replicates of C and U sRNA libraries were prepared combining the protocols of mDOT-seq and NEXTflex Small RNA-Seq Kit v3 (PerkinElmer), while for N libraries RNA was processed only according to the later one. All libraries were pooled together and sequenced at Lexogen facilities using NextSeq 500/550 Mid Output Kit to obtain 2 x 75 paired-end reads.