This document provides an overview of multiple sequence alignment (MSA). MSA is used to align biological sequences, such as DNA, RNA, or proteins, to find similarities and differences between sequences. The document outlines the goal of MSA as finding structural, functional, and evolutionary relationships. It describes the general considerations and steps of MSA, including pairwise comparison of sequences, cluster analysis to generate a hierarchy, and progressive alignment. Common MSA software and applications are also summarized.

1. MULTIPLE SEQUENCE ALIGNMENT
Name – Swati Kumari
M.Sc. Bioinformatics
Roll No – 22
Central University of
Bihar, Patna

2. CONTENTS
 Introduction
 Goal of MSA
 General Considerations for MSA
 Steps of MSA
 Diagrammatic representation of steps of
MSA
 Applications of MSA
 Software for MSA
 References

3. INTRODUCTION
 A Multiple Sequence Alignment (MSA) is
alignment of biological sequences to find the
similarity or dissimilarity between
sequences, where the number of sequences
are more than two.
 One of most essential tools in molecular
biology applied for both amino acids and
nucleotides (DNA,RNA).

4. Goal of MSA
 To find the Similarities based on Nucleotide
or Amino Acid.
 To find the Functional similarities.
 To find the Structural similarities.
 To find the Evolutionary relationship.

5. General Considerations for MSA
 The more number of sequences to align gives the
better result.
 Only 40% of similarities between two amino
acid sequences shows they are close in structure.
 Sub­group of n number of sequences should be
pre­aligned separately, and one member of each
sub­group should be included in the final
multiple alignment.

6. Steps of MSA
 Compare all sequences pairwise.
 Perform cluster analysis on the pairwise data to
generate a hierarchy for alignment. This may be
in the form of a binary tree.
 Build the multiple alignment by first aligning the
most similar pair of sequences, then the next
most similar pair and so on.

7. diagrammatic representation of steps of MSA

8. Applications of MSA
 Identify new protein or gene families.
 Determining the relation between the aligned
protein.
 Development of a genetic representation of
protein family.
 In practical analysis ­
 Mutant analysis
 Identify conserved primer binding site.

9.  Designing experiment to test and modify the
function of the aligned sequences –
 Identify amino acids crucial for function.
 Locating non­ conserved region / useful or tag
insertion.
 Designing degenerate primer for Polymerase
Chain Reaction (PCR).

10. Software for MSA
 Clustal­W (Command Line Version) ­
 the famous multiple alignment program for
nucleic acid and protein sequences.
 Clustal­X (Graphical Line Version) –
 provides a window­based user interface to the
Clustal­W multiple alignment program.
 MUSCLE (multiple sequence comparison
by log­expectation) –
 more accurate than T­Coffee, faster than
Clustal­W.

11.  T­Coffee (Tree­ based Consistency Objective
Function For alignment Evolution)
 Main characteristics is allow to combine result
obtained with several alignment method.
 Kalign ­
 It is very fast MSA tool that concentrates on
local region suitable for large alignment.

12. References
 http://www.bioinformatics.org/wiki/Multiple_seq
uence_alignment
 http://
en.wikipedia.org/wiki/Multiple_sequence_alignm
ent
 www.clustal.org/
 http://www.ebi.ac.uk/Tools/msa/
 Class lecture

13. THANK YOU