Human cDNA filter Laboratory for Cancer Research, Rutgers University. The arrays contains 10,692 PCR amplifications of human cDNAs from a collection of EST clones purchased from Research Genetics. 10,692 EST clones were selected from a Research Genetics EST collection focusing on plates that contained mostly known genes. There are approximately 7,000 ESTs corresponding to known genes, and the remainder are ESTs either with homology to known genes or are anonymous. cDNA inserts of the EST clones were amplified by PCR in a 96-well format (MJ Research, Boston, Massachusetts) using M13 forward (5'-CGTTGTAAAACGACGGCCAGT-3') and reverse primers (5'-TTTCACACAGGAAACAGCTATGAC-3'). The 10,692 PCR products were transferred onto 3x10 in Hybond N+ nylon membranes (Amersham Pharmacia Biotech) using the GeneMachines microarrayer. After spotting the arrays were cross-linked by UV. To assure even quality between subsequent rounds of hybridization, cDNA arrays were pre-stripped before their first use. To check for filter quality, human COT-1 DNA hybridizations were carried out. Keywords = membrane, filter
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