title: Custom NimbleGen 12-plex Bufo marinus, Bufo boreas, Xenopus tropicalis Oligo Expression Array
status: Public on Apr 18 2017
contact.city: Berkeley
contact.first: Thomas
contact.last: Poorten
contact.email: tom.poorten@gmail.com
contact.state: CA
contact.address: 54 Mulford Hall
contact.department: Environmental Science, Policy, and Management
contact.country: USA
contact.institute: UC Berkeley
organism: Xenopus tropicalis
  Rhinella marina
  Anaxyrus boreas
description: The custom Nimblegen microarray design included 135,200 60-bp probes (excluding control probes) targeting 31,367 transcript contigs from Bufo marinus, Bufo boreas, and model species Xenopus tropicalis, with 4 probes per probe-set. Probe-sets were defined as all probes that mapped to a single transcript. We used the 12-plex microarray platform (12 arrays per glass slide). Annotation was performed using the blat program to identify Bufo contigs with high sequence identity to X. tropicalis transcripts (Ensembl assembly v4.2). We ran blat with translated Bufo contigs and translated X. tropicalis transcripts (parameters: -q=dnax –t=dnax) in order to find orthologous sequences at the protein level, which is more conserved between divergent species. We parsed blat hits to remove short hits (<100 bp aligned to query) and low scoring hits (Expectation value >1.0e-12). We used the Ensembl Gene Identifier (e.g. ENSXETG00000XXXXXX) for the blat hit with longest alignment to the query as the final annotation call. We then imported annotation metadata with the biomart.org pipeline: Ensembl gene name and description, Gene Ontology (GO) terms, Ensembl protein families, Interpro protein domains. Next, we used the blast2go pipeline to annotate Bufo contigs without GO term metadata (6328 contigs). We used the blastx method with the NCBI nonredundant protein database to identify blast hits, and proceeded with blast2go pipeline with default settings.
manufacture_protocol: See manufacturer's website