A small CRISPR/Cas-associated RNA regulates ferrous iron transport in Neisseria meningitidis

In this study the RNA targetome of the endonuclease Cas9, the trans-activating crRNA (tracrRNA) and a CRISPR-associated small RNA (scaRNA/NMnc0040) was determined in Neisseria meninigitidis serogroup C strain 8013 by RNA-seq. To uncover the RNA targetome of Cas9, tracrRNA and scaRNA of Neisseria meningitidis (N.meninigitidis), bacterial samples were collected of three biological replicates of N.meninigitidis strain 8013 wt and cas9, tracrRNA and scaRNA deletion in which the CDS of Cas9 and the sequences of tracrRNA and scaRNA was replaced by an antibiotics cassette. Bacterial cultures grown to OD600nm of 0.5 in cell culture medium (EMEM+++) were snapfrozen in liquid nitrogen prior to total RNA-extraction with the hot phenol method and DNase digestion.