title: Fred Hutchinson Cancer Research Center human 4.3K BAC array
status: Public on Aug 17 2008
contact.city: Seattle
contact.first: Ilona
contact.middle: N
contact.last: Holcomb
contact.email: iholcomb@u.washington.edu
contact.state: WA
contact.address: 1100 Fairview Ave N
contact.department: Human Biology
contact.country: USA
contact.institute: Fred Hutchinson Cancer Research Center
organism: Homo sapiens
manufacture_protocol: This array consists of 4342 clones that are placed at a single location on the May 2004 draft assembly. 99% of map locations have been FISH verified. The clones have a median spacing of 402 kb across the chromosomes, excluding the repetitive regions around the centromeres, the short arms of the acrocentric chromosomes, and the long arm of the Y chromosome.
  To construct the array, BAC DNA was extracted from 1.5-mL cultures with a modified Qiagen R.E.A.L. Prep96 plasmid kit (Qiagen). Purified BAC DNA was then amplified with ligation-mediated PCR (as indicated in Klein C, Schmidt-Kittler O, Schardt J, Pantel K, Speicher M, Riethmuller G Comparative genomic hybridization, loss of heterozygosity, and DNA sequence analysis of single cells. Proc Natl Acad Sci USA 1999;96:4494-9.) and cleaned with the Multiscreen-PCR filtration system (Millipore) on a Biomek FX automated liquid handling system (Beckman Coulter, Fullerton, CA). Purified products were transferred to 384-well plates, air-dried, and then resuspended in 3x SSC. Each amplified/purified BAC was then mechanically spotted in triplicate onto poly-L-lysine-coated slides using an Omnigrid arrayer (GeneMachines, San Carlos, CA).