Sequence Similarity Search, Multiple Sequence Alignment, Model Selection, Distance Matrix and Phylogeny Reconstruction

Abstract

This is a generic sequence analysis protocol suitable for plant and algal phylogeographic studies. Generated sequences from bidirectional Sanger sequencers are first assembled using Geneious. Sequence assembly is then trimmed and similarity search is conducted using BLASTn within Geneious. BLAST hits and other target taxa are selected and multiple sequence alignment is constructed. The alignment is then refined by checking using eye and exported as .fasta. Using MEGA, best-fitting nucleic acid substitution models will be calculated in MLModelTest. Model with lowest BIC score is selected and used for further phylogenetic analysis using MEGA or Geneious, which include distance matrix construction, phylogeny reconstruction using ML and BI.