bioinformatics for nucleotide sequence analysis

BIOINFORMATICS FOR NUCLEOTIDE

SEQUENCE ANALYSIS

BIOC 2112125-05-2015

Home page of NCBI

http://www.ncbi.nlm.nih.gov/

What can we do with NCBI ?

To find similar sequences – Sequence AlignmentTo find the relatedness of a gene or protein (do they have a common ancestor?)To study mutationsPrimer designing

WHAT IS BLAST ? Basic Local Alignment Search Tool A method for rapid searching of nucleotide

and protein databases Offers both sensitivity and speed Fastest and most frequently used sequence

alignment tool

Align 2 or more sequencesPrimer blast

BLAST ACCESS NCBI BLAST http://www.ncbi.nlm.nih.gov/BLAST/ Canadian Bioinformatics Resource BLAST http://cbr-rbc.nrc-cnrc.gc.ca/blast/ European Bioinformatics Institute BLAST http://www.ebi.ac.uk/blastall/ http://www.ebi.ac.uk/blast2/

BLAST - PROGRAMS Blastp - Compares an amino acid query sequence against a

protein sequence database.

Blastn - Compares a nucleotide query sequence against a nucleotide sequence database.

Blastx - Compares a nucleotide query sequence translated in all reading frames against a protein sequence database.

Tblastn - Compares a protein query sequence against a nucleotide sequence database.

Accession Number - NM_008862

TCGAAATAACGCGTGTTCTCAACGCGGTCGCGCAGATGCCTTTGCTCATC AGATGCGACCGCAACCACGTCCGCCGCCTTGTTCGCCGTCCCCGTGCCTC AACCACCACCACGGTGTCGTCTTCCCCGAACGCGTCCCGGTCAGCCAGCC TCCACGCGCCGCGCGCGCGGAGTGCCCATTCGGGCCGCAGCTGCGACGGT GCCGCTCAGATTCTGTGTGGCAGGCGCGTGTTGGAGTCTAAA

Q1 : Find the protein coded by the following nucleotide sequence.

http://www.ncbi.nlm.nih.gov/

BLAST StatisticsThe Score and the E-value (E) are particularly helpful in making interpretations.

Score - a good measure of the quality of an alignment.E-value - expectation value is a good measure of the

significance of the alignment.

The E-value is the number of different alignments, with scoresequivalent to or better than S, that are expected to occur in a database search by chance.

Significant the alignmentE value

Toll-like receptor 3 - human - NM_003265 - mouse - NM_126166 - zebra fish - NM_001013269

Q2 : Find the relatedness of the following receptors.

http://www.ncbi.nlm.nih.gov/

Crassostrea gigas heat shock protein70

Q3 : Design primers for the following gene.

http://www.ncbi.nlm.nih.gov/

Q4 : Determine the fragment size of the following PCR product.

TubA1 U12589Alpha tubulin gene of Forward 5’ GGAAACGCCTGCTGGGAGReverse 5’ AACAGTTGGAGGCTGATAAT

http://www.ncbi.nlm.nih.gov/

CONCLUSIONS BLAST is one of the most important program in

bioinformatics (maybe all of biology) BLAST is based on sound statistical principles

(key to its speed and sensitivity) A basic understanding of its principles is key for

using/interpreting BLAST output