Direct detection of Staphylococcus aureus mRNA using a flow through microarray.
The direct detection of mRNAs from bacterial cultures on a DNA array without amplification and labelling would greatly extend the range of applications suitable for microarray analysis. Here we describe the direct detection of 23S rRNA and seven mRNA species from total Staphylococcus aureus RNA prepared using commercially available RNA purification columns followed by fluorescent detection on a flow through microarray. RNA hybridisation was detected using paired secondary labelled probes directly 5' and 3' to immobilised 60 mers. In this way, we were able to detect the effect of 30-min exposure to antimicrobials on mRNA levels within 3 h after column purification of total RNA without the need for enzymatic manipulation. Specifically the expression of mecA was confirmed in a highly resistant strain and induction of katA and ile-tRNA synthetase genes after exposure to mupirocin could be detected.
Anthony
R M
RM
KIT Biomedical Research, Meibergdreef 39, 1105 AZ Amsterdam, The Netherlands. r.anthony@kit.nl
Schuitema
A R J
AR
Oskam
L
L
Klatser
P R
PR
D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D012338:RNA, Ribosomal, 23S / Q000737:chemistry / Q000235:genetics; D013211:Staphylococcus aureus / Q000235:genetics*
D012329:RNA, Bacterial; D012333:RNA, Messenger; D012338:RNA, Ribosomal, 23S