title: Genomic Research Laboratory Rickettsia conorii 0.7 k
status: Public on Jul 06 2006
contact.city: Geneva
contact.first: FRANCOIS
contact.last: Patrice
contact.email: patrice.francois@genomic.ch
contact.state: Ge 4
contact.address: Rue Gabrielle-Perret-Gentil, 4
contact.department: Service of Infectious Diseases
contact.country: Switzerland
contact.institute: Genomic Research Laboratory
contact.phone: +41 (0)22 372 93 37
organism: Rickettsia conorii
manufacture_protocol: A microarray including 129 targets specific for R. conorii was constructed by PCR amplification. These targets include 82 ORFs representing about 6% of R. conorii genome, and 47 intergenic spacers. Briefly, a specific amplification of selected targets was carried out usi printed onto glass slides (Corning® Gaps II Coated slide) by using a SDng oligonucleotide primers containing an universal sequences at the 5’-end (5’-CGACGCCCGCTGATA-3’ for forward / 5’-GTCCGGGAGCCATG-3’ for reverse). Following purification (MultiScreen PCR96 Filter Plate, Millipore, Billerica, MA, USA), amplicons (QIAquick 96-well purification kit ; QIAGEN, Courtaboeuf, France) were sequenced (3100 Genetic Analyzer, Applied Biosystems, Courtaboeuf, France) to check for the identity of amplified samples and processed in a second PCR run by using universal primers. The amplified DNA fragments were thenDC-2 robot (Bio-Rad, Marnes-la-Coquette, France). The spotted slides were cross linked by using a UV Stratalinker (Stratagene, La Jolla, CA, USA) each at a total energy of 250 mJ and stored in a dry environment at room temperature for at least 12 hours. The quality of printing (spot morphology and intensity) was checked by Syto-9 staining (Molecular Probes, Eugene, OR).