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Practical: Multiple Sequence Alignment and building a Phylogenetic Tree using ClustalW


For building a phylogenetic tree, you need more than 2 sequences for both multiple sequence alignment and building a tree using ClustalW. 
Phylogenetic Tree in bioinformatics represents the evolutionary relationship between species. 

 Follow these steps to build a tree:

 These steps are for building a phylogenetic tree for p53 of 4 different species (Homo sapiens, Mus musculus, Rattus norvegiccus, Danio rario). (You can consider the protein or dna of your interest to find the relationship between any number of species). 

 1) Go to NCBI homepage and select Nucleotide. And type in the search box "p53" and you will get number of hits for this query. And a number of entries for p53 in different species will be displayed.

 2) Consider 4 different species (example: homo sapiens, mus musculus, rattus norvegiccus, danio rario). and retrieve FASTA format sequneces for those 4 species and paste them in a word file one by one. 


 3) Open https://www.ebi.ac.uk/Tools/msa/clustalw2/ 
     (Since Clustalw2 has retired, you need to go to Clustal Omega for multiple sequence alignment and tree). and click on multiple sequence alignment first. 

 4) Click on Clustal Omega and the query box will be displayed. 

 5) Copy the FASTA format sequences and paste them in that query box. and click RUN. 

 6) After a while, the results for multiple sequence alignment will be displayed. 

 7) Go through the multiple sequence alignment file for better view and understanding of how nucleotides are related. 

 8) Now, copy the job ID, which looks like 
clustalo-I20190205-151110-0669-40920939-p1m

 9) Copy this Job ID and paste it on wordpad for reference. 

 10) In the same webpage, there is an option for Phylogenetic Tree next to the multiple alignment. 

 11) Click on Phylogenetic Tree and The tree will be displayed. 

 12) The phylogenetic tree will be displayed as shown in the image below. The built tree represents the evolutionary relationship between species for p53. 


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