Computational protocol: Runx Family Genes in a Cartilaginous Fish, the Elephant Shark (Callorhinchus milii)

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Protocol publication

[…] The elephant shark 1.4× coverage sequence assembly (http://esharkgenome.imcb.a-star.edu.sg/) was searched with human RUNX1, RUNX2, RUNX3 and RUNXβ protein sequences using ‘TBLASTN’ algorithm. The following contigs that contained fragments of various Runx genes were identified in the assembly: Runx1- AAVX01052077, AAVX01261782, AVX01381831 and AAVX01569464; Runx2 - AAVX01175228, AAVX01545426, AAVX01631487, AAVX01048938, AAVX01307838 and AAVX01048937; Runx3 - AAVX01039927, AAVX01288244, AAVX01157201 and AAVX01598430 and Runxb- AAVX01192354, AAVX01224391 and AAVX01471917. [...] The protein sequences for human and other chordate Runx genes were retrieved from the National Center for Biotechnology Information (NCBI) database. Multiple sequence alignments were performed by using ClustalW. For phylogenetic analysis, gaps in the alignments were removed using Gblocks Server (ver. 0.91b) with default parameters . The phylogenetic analysis was performed using MrBayes 3.2, employing Dayhoff+G as a substitution model and running 4 chains for 1,000,000 generations. Trees were sampled every 100 generations and according to a saturation curve of likelihood values, the first 2500 trees were discarded as burn-in. The Runt gene from Branchiostoma floridae was used as the outgroup. [...] Genomic sequences of the Runx gene loci for the following species were extracted from Ensembl release 73 : human (GRCh37 assembly, February 2009), mouse (GRCm38, December 2011), dog (CanFam3.1, September 2011), opossum (MonDom5, October 2006), chicken (GalGal4, November 2011), lizard (AnoCar2.0, May 2010), frog (JGI 4.2, November 2009), zebrafish (Zv9, July 2010) and fugu (FUGU5, October 2011). Repetitive sequences were masked using CENSOR at default settings . Multiple alignments of Runx gene loci sequences were generated using the global alignment program MLAGAN (http://genome.lbl.gov/vista/index.shtml) with human as the reference sequence. CNEs were predicted using a cutoff of ≥65% identity across 50-bp windows and visualized using VISTA (http://genome.lbl.gov/vista/index.shtml). […]

Pipeline specifications

Software tools Clustal W, Gblocks, MrBayes, LAGAN
Applications Phylogenetics, Nucleotide sequence alignment
Organisms Danio rerio, Callorhinchus milii, Homo sapiens