Computational protocol: Splicing Variants of SERPINA1 Gene in Ovine Milk: Characterization of cDNA and Identification of Polymorphisms

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Protocol publication

[…] Whole mammalian genome scanning was done to identify the homologous regions of the full length sheep SERPINA1 cDNA and gene using Basic Local Alignment Search Tool (http://www.ncbi.nlm.nih.gov/BLAST/). Sequence data were edited, translated and aligned using the free software Bioalign 4.0.6 (http://en.bio-soft.net/dna/BioLign.html). The open reading frame (ORF) of the full-length AAT cDNA was determined by ORF Finder at NCBI (www.ncbi.nlm.nih.gov/gorf/).To identify SNPs with potential impact on splicing of SERPINA1 gene, mutant and wild sequences were analyzed with the Human Splicing Finder software (http://139.124.156.135:2300/), which includes several matrices to analyze splice sites and splicing silencers and enhancers.To determine the potential deleterious effect of amino acid changes on protein function we used the SIFT (http://blocks.fhcrc.org/sift/SIFT.html) software. This software uses the protein sequence similarity of different species and the characteristics of amino acids (structure, polar/no polar, basic/acid) to calculate the probability of a deleterious effect of specific amino acid variants. Scores lower than 0.05 suggest a potential not tolerated amino acid substitution and a potential influence on protein function.To search for homology of the predicted protein sequence with other species the BLASTP software was used (http://www.ncbi.nlm.nih.gov/BLAST/). We aligned the AAT protein sequences of different organisms with MEGA5 software (http://www.megasoftware.net/) [] to examine the evolutionary conservation of RCL motifs. […]

Pipeline specifications

Software tools BLASTN, BioLign, Open Reading Frame Finder, HSF, SIFT, BLASTP, MEGA
Applications WGS analysis, Nucleotide sequence alignment
Organisms Ovis aries, Homo sapiens, Bos taurus