title: Marinebiotechnology laboratory Haematococcus pluvialis 1k cDNA microarray platform
status: Public on Sep 22 2006
contact.city: InCheon
contact.first: HyunSuk
contact.last: Eom
contact.email: eomyanga@hotmail.com
contact.state: Nam-gu
contact.address: 253
contact.country: South Korea
contact.institute: Inha University
organism: Haematococcus lacustris
manufacture_protocol: We constructed the cDNA microarrays from the Haematococcus pluvialis cDNA library (Eom et al., 2005). One thousand selected expressed sequence tags (ESTs) from each clone, including housekeeping genes like ribosomal binding proteins, were amplified by PCR using T3/T7 primers or M13-20/M13 reverse primers and recovered by a MultiScreen PCR Filter Plate (Millipore, Billerica, MA, USA). Each cDNA chip was designed with 965 successfully amplified PCR products from the cDNA library. Additionally, several control clones from the Saccharomyces cerevisiae intergeneic sequence and the Haematococcus pluvialis housekeeping genes were included. Microarrays were produced using a fully automated system by robotic operation at Digital Genomics (Seoul, South Korea). The cDNA PCR products were spotted onto amine-coated GAPSII slides (Corning Inc., Corning, NY, USA) using MG II (Biorobotics Inc., Woburn, MA, USA) with a 240 um pitch between spots as a twin format. There were 18 x 18 spots per block, and the spot diameter was 100~140 um. To evaluate fabrication, a cDNA microarray was stained with Syto61 (Molecular Probes, Eugene, OR, USA).