title: CRIBI pig mRNA expression array [platform 2]
status: Public on Mar 31 2014
contact.city: Padova
contact.first: Gerolamo
contact.last: Lanfranchi
contact.email: stefano.cagnin@unipd.it
contact.address: Via U. Bassi, 58/B
contact.department: CRIBI - Biotechnology Center and Biology Department
contact.country: Italy
contact.institute: University of Padova
contact.phone: +39-0498276219
organism: Sus scrofa
description: Concerning mRNAs Ensembl transcripts (Ver. 56) and UniGene (Ver. 38) sequences were used to produce a pig dedicated microarray platform to analyze mRNAs gene expression. Based on sequence similarity we rejected UniGene features that overlapped more than 40% with an Ensembl transcript. After this filter we obtained 40,267 UniGene clusters and 19,603 Ensembl transcripts (protein coding + pseudogenes + retrotransposed elements). Starting from these sequences, the microarray probes were computed using 6 different algorithms. We obtained probes with different specificity and different distances from the mRNA 3 -end. The two best probes for every sequence, basing on the reliability of the prediction algorithm and their vicinity to the 3 -end were experimentally tested for their specificity and performance (GEO: ..). For any transcript with a replicated probe, we selected the probe that was the most responsive and specific in a hybridization trial performed with a pool of mRNAs from 20tissues. The specificity was based on the intensity fluorescence, as suggested by Kronick (Kronick 2004). Replicated probes with higher intensity fluorescence were considered the most responsive for the specific gene. The resulting whole genome microarray, here used for the gene expression analysis, was composed by 17,048 replicated probes and 963 not replicated specific for the Ensembl transcripts, 11,363 replicated probes specific for the UniGene clusters of length comprised between 778 nt and 1,348 nt (Figure SM1), and 28,790 single probes specific for the remaining UniGene clusters. All probes mapped proximally to the 3 end of the target sequences (Figure SM2 A and SM2 B): 98.2% of the probes for the Ensembl transcripts mapped within 2,000 nt from their 3 end while the percentage for UniGene clusters reached 99.9%. 114 UniGene clusters and 1,592 Ensembl transcripts were not identified by specific probes.
manufacture_protocol: 90K Combimatrix microarray were synthesized using the Combimatrix synthesizer (Combimatrix) that allows in-situ synthesis of oligonucleotide probes through the phosphoramidites chemistry.