ena-EXPERIMENT-RUDER BOSCOVIC INSTITUTE-03-04-2019-19:24:31:160-29
Illumina MiSeq paired end sequencing
cDNA sequencin of 16S rRNA gene from the chemocline, monimolimnion and sediment of meromictic lake
unspecified
Total RNA was treated with Turbo DNase (2 U/μl, Ambion) at 37 °C for 30 min. . The Turbo DNase was inactivated at room temperature for 5 min.To the RNA, 1.5 μl of a 10-mM deoxyribonucleoside triphosphate (dNTP) mix (New England Biolabs), 0.75 μl of a 50 mM MgCl₂ (Sigma Aldrich, USA) and 2.25 μl of random primers (60 μM) (New England Biolabs) was added, heated  at 75 °C for 4 min. and chilled on ice. 3 μl of 10× first-strand buffer [500 mM Tris–HCl (pH 8.3), 750 mM KCl, 30 mM MgCl2, 50 mM DTT], 1 μl of SUPERase-In RNase Inhibitor (10 U/μl, New England Biolabs), 1.5 μl of Moloney Murine Leukemia Virus–Reverse Transcriptase (M-MuLV) (200 U/μl, New England Biolabs) were added to yield a final reaction volume of 25 μl.
RUDER BOSCOVIC INSTITUTE
ena-EXPERIMENT-RUDER BOSCOVIC INSTITUTE-03-04-2019-19:24:31:160-29