Published by Pratik Umesh Parikh Master of Pharmacy (Pharmaceutical Biotechnology)

1. Sequence
Analysis
PRATIK UMESH PARIKH
MASTER OF PHARMACY
(PHARMACEUTICAL BIOTECHNOLOGY)

2. Contents :
❖Introduction
❖DNA & protein sequencing
❖Sequence Assembly
❖Sequence Alignment
❖Multiple Sequence Alignment
❖Demonstration of Protein seqeuncing
❖Applications
❖Reference

3. Sequence analysis :
Sequence Analysis is the process of subjecting a DNA, RNA or peptide sequence
to any of a wide range of analytical methods to understand its features, function,
structure, or evolution.
It Includes :
Sequencing: Sequence Assembly ANALYSIS
Alignment: Searching (in Databases)

4. DNA & Protein Sequencing :
DNA sequencing :
DNA sequencing is the process of determining the precise order of nucleotides
or order of the four bases—adenine, guanine, cytosine, and thymine, in a strand
of DNA.
Protein sequencing :
Protein sequencing is a technique to determine the amino acid sequence of a
protein, as well as which conformation the protein adopts and the extent to
which it is complexed with any non-peptide molecules.

5. Sequence assembly :
Sequence assembly refers to the reconstruction of a DNA sequence by aligning
and merging small DNA fragments.
It is an integral part of modern DNA sequencing :
(1) cutting the DNA into small pieces,
(2) reading the small fragments,
(3) reconstituting the original DNA by merging the information on various
fragment.

6. Sequence Alignment :
Sequence Alignment is a way of arranging the sequences of DNA, RNA, or protein
to identify regions of similarity that may be a consequence of functional,
structural, or evolutionary relationships between the sequences.
It involves the identification of the correct location of deletions and insertions
that have occurred in either of the two lineages since the divergence from a
common ancestor.
TYPES :
On the basis of number of comparing sequencing strand, it is of two types:
➢Pair-wise Alignment
➢Multiple Sequence Alignment

7. Multiple sequence Alignment :
Multiple sequence alignment (MSA) may refer to the process or the result of sequence
alignment of three or more biological sequences, generally protein, DNA, or RNA.
Tools Used : ClustalW, ClustalX, BLAST, FASTA, Phyre2

8. Demonstration of Protein Sequencing :
1. Find the human prion protein in NCBI and retrieve the sequence and save it in computer.
2. Run BLAST to find similar sequences and retrieve some similar sequences in FASTA format.
3. Look for the presence of conserved domain (If any).
4. Go for the multiple sequence alignment with 10 of the retrieved sequences.
5. Construct the phylogenetic tree with the sequence alignment.
6. Predict the secondary structure of the human prion protein.

9. Find the human prion protein in NCBI and retrieve the sequence and save
it in computer :
https://www.ncbi.nlm.nih.gov/

10. Run BLAST to find similar sequences and retrieve some similar sequences in FASTA
format.
https://blast.ncbi.nlm.nih.gov/Blast.cgi

11. Go for the multiple sequence alignment with 10 of the retrieved
sequences.
https://www.ebi.ac.uk/Tools/msa/clustalo/

12. Predict the secondary structure & 3D view of the human prion protein
https://swissmodel.expasy.org/
http://www.sbg.bio.ic.ac.uk/
phyre2/html/page.cgi?id=ind
ex

13. Applications :
➢ The comparison of sequences in order to find similarity, often to infer if they are related
(homologous).
➢ Identification of intrinsic features of the sequence such as active sites, post translational
modification sites, gene-structures, distributions of introns and exons.
➢ Identification of sequence differences and variations such as point mutations and single
nucleotide polymorphism (SNP) in order to get the genetic marker.
➢ Revealing the evolution and genetic diversity of sequences and organisms
➢ Identification of molecular structure from sequence alone.
➢ Genetic diseases

14. Reference :
1. https://www.slideshare.net/PrashantTripathi59/sequence-analysis-71940516
2. https://www.ncbi.nlm.nih.gov/
3. https://blast.ncbi.nlm.nih.gov/Blast.cgi
4. https://www.ebi.ac.uk/Tools/msa/clustalo/
5. https://swissmodel.expasy.org/
6. http://www.sbg.bio.ic.ac.uk/phyre2/html/page.cgi?id=index