title: Cavalieri-Yeast-ORF-7k Array
status: Public on Dec 09 2009
contact.city: Torino
contact.first: Irene
contact.last: Stefanini
contact.email: stefanini.irene@gmail.com
contact.address: via Accademia Albertina
contact.country: Italy
contact.institute: University of Turin
organism: Saccharomyces cerevisiae
manufacture_protocol: Yeast microarrays were constructed using the amplifying to levels required for preparation of DNA microarrays by PCR (27) a set of clones containing 6218 verified ORFs of the yeast genome obtained from Research Genetics (Huntsville, AL). Some of the longer ORFs were amplified with the GIBCO/BRL Elongase Amplification Kit (Life Technologies, Rockville, MD), using 40 cycles of 1 min at 95 °C denaturation, 1 min at 55 °C annealing, and 10 min at 68 °C elongation. We obtained an amplified product confirmed by agarose gel electrophoresis for 98% of the ORFs. The amplified DNA was precipitated with isopropanol, washed with 70% EtOH, and resuspended in 25 μl Micro Spotting Solution (Telechem, Sunnyvale, CA). The DNA was spotted on CMT-GAPS amino-silane-coated glass slides (Corning), using a microarraying robot with a 16-pin head constructed from a design by Patrick O. Brown (http://cmgm.stanford.edu/pbrown/).