title: UniMuenster-HomoSapiens-30K-Promoter_v1
status: Public on Aug 22 2010
contact.city: New York
contact.first: Hans-Ulrich
contact.last: Klein
contact.email: h.klein@uni-muenster.de
contact.state: NY
contact.address: 622 W 168th St
contact.department: Neurology
contact.country: USA
contact.institute: Columbia University Medical Center
organism: Homo sapiens
description: Human putative promoter sequences of all known genes and sca/miRNA genes were downloaded from UCSC Golden Path Database based on NCBI Build 35. The putative promoter sequences were defined as 2,000 bases upstream and 1,000 bases downstream from transcription start site. Sequences were repeat-masked. Probes were designed from the sense strand of the non-repeat-masked portions of the promoter regions, with these probe restrictions: 1) Length: 46 – 55 bases 2) Tm: 69 – 79 °C 3) Creatable by NimbleGen in 148-rounds of A, C, G, T. 4) Probe must not contain simple repeats. 5) Probe must not have tendency to form secondary structure. A second set of miRNA probes was created with a slightly looser length restriction of 40 – 60 bases, to be used for hard-to-cover miRNAs. The number of probes was reduced by choosing the closest probe to every position in the promoter that was a multiple of 25, thus spacing the probes roughly 25 bases apart. Probe specificity was screened by aligning them to the human genome using Blat. Probes that had imperfect matches in the genome of 30 bases or more were thrown out, as were probes that had more than 2 exact matches. Up to 2 upstream probes and 1 downstream probe were selected for each promoter region, with an attempt made at sharing probes among overlapping promoter regions.  Priority was given to probes that contained the degenerate sequence RCG or CGY, and that were found within 1000 bases upstream and 500 bases downstream of the start of transcription. Genes with known functions were prioritized for oligo synthesis. Most genes have three representative oligos. After addition of controls, the final array contained the following numbers of spots: 288 blank spots, 366 spotting buffer spots, 192 GFP spots, 176 Salmonella spots and 30178 regular spots.
manufacture_protocol: Oligonucleotides were synthesized with 5´ aminolinker by Operon (Cologne, Germany) and spotted onto Nexterion slides.