title: PBI-NRC_Brassica 10K V1.0
status: Public on May 31 2009
contact.city: Saskatoon
contact.first: Jitao
contact.last: Zou
contact.email: jitao.zou@nrc-cnrc.gc.ca
contact.state: Saskatchewan
contact.address: 110 Gymnasium Place
contact.country: Canada
contact.web_link: http://pbi-ibp.nrc-cnrc.gc.ca/ENGLISH/research-officers/zou-j.htm
contact.institute: Plant Biotechnology Institute, National Research Council Canada
contact.phone: 1-306-975-5583
organism: Brassica napus
description: A set of 10,642 PCR primer pairs were designed and corresponding amplicons were produced for preparation of a targeted seed cDNA microarray. cDNA amplicons were printed on Corning UltraGAPS slides with the Chipwriter Pro microarray printer (Bio-Rad). The microarray consists of 12 row × 4 column sub-arrays, and each sub-array has 23 row × 22 column (506) spots. Spots were printed in duplicate side-by-side by columns, and the total number of spots on the microarray is 24288, of which 22,656 spots are cDNA probes. Controls are printed at the beginning and the end of each sub-array. The distance between spot is 185 µm and the spot diameter is 145 µm.
manufacture_protocol: Protocol for cDNA array fabrication
  1)	Amplify the insertions with a GeneAmp PCR System 9700 (Applied Biosystems, ABI) using AmpliTaq (ABI);
  2)	Purify the PCR products using the Biomek FX (Bechman/Coulter) and check the quality through agrose gel electrophoresis;
  3)	Amplicons were consolidated into 384-well plates, lyophilized, and resuspended in 10 µl of 50% DMSO to a final concentration of 250 ng/µl.
  4)	The Stratagene SpotReport Alien cDNA Array Validation kit and an in-house Candida albicans ECE1 gene (L17087) were ready in a dilution series. The buffer/blank control was 50% DMSO;
  5)	Printing the microarray on Corning UltraGAPSTM slides using the VersArray ChipwriterTM Pro microarray printer (Bio-Rad). SMP3 Microspot quill pins (TeleChem) in a 48-pin tool to deposit ~7 nL per spot onto the slide;