Computational protocol: Conserved developmental expression of Fezf in chordates and Drosophila and the origin of the Zona Limitans Intrathalamica (ZLI) brain organizer

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Protocol publication

[…] Using the previously described vertebrate Fezf genes as queries, we performed tBLASTN and/or BLASTP searches against the genomes of Branchiostoma floridae JGI v1.0, Trichoplax adhaerens Grell-BS-1999 v1.0, Nematostella vectensis JGI v1.0, Ciona intestinalis JGI v2.0, Daphnia pulex JGI v1.0, Lottia gigantea JGI v1.0 and Capitella teleta JGI v1.0, using the JGI website (http://genome.jgi-psf.org/euk_home.html) and of Strongylocentrotus purpuratus Build 2.1, Tribolium castaneum Build 2.1, Nasonia vitripennis Build 1.1, Drosophila melanogaster Build Fb5.3, Homo sapiens Build GRCh37, Mus musculus Build 37.1, Danio rerio Build Zv8, using the NCBI website (http://www.ncbi.nlm.nih.gov/blast/Blast.cgi). For Saccoglossus kowalevskii we performed a tBLASTN search against the traces at NCBI and then manually assembled the genomic locus.We then downloaded each corresponding genomic region and build different gene models using GenomeScan [] and GeneWise2 [] software as necessary. We compared these predictions with expressed sequence tags and existent gene models when available. Annotation and comparison of intron positions and phases across zinc finger domains was performed as previously described [,].The amino acid sequences for the zinc finger domains were aligned using ClustalW [] and the resulting alignment was manually curated. Phylogenetic trees were then generated by the Bayesian method, using the software MrBayes 3.1.2 [,], with the model Dayhoff+Gamma, recommended by ProtTest 1.4 [-], under the Akaike information and the Bayesian information criterions. Two independent runs were performed, each with four chains. For convention, convergence was reached when the value for the standard deviation of split frequencies stayed below 0.01. Burn-in was determined by plotting parameters across all runs for a given analysis: all trees before stationarity and convergence were discarded, and consensus trees were calculated for the remaining trees (from at least 1,000,000 generations).Fezf box consensus was decided by the program Sequence Logo online (http://genome.tugraz.at/Logo/) using a multiple alignment for all studied species containing a Fezf box. […]

Pipeline specifications

Software tools TBLASTN, BLASTP, GenomeScan, GeneWise, Clustal W, MrBayes, ProtTest
Applications Phylogenetics, Amino acid sequence alignment
Organisms Drosophila melanogaster, Branchiostoma lanceolatum