S.Prasanth Kumar, S.Prasanth Kumar, BioinformaticianBioinformatician

Identification of Disease GenesIdentification of Disease Genes

Pharmacogenomics & Drug Design

S.Prasanth Kumar, S.Prasanth Kumar, BioinformaticianBioinformatician

S.Prasanth Kumar Dept. of Bioinformatics Applied Botany Centre (ABC) Gujarat University, Ahmedabad, INDIA

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From Scratch

Patient New Symptom

Phenotype

Genotype

Molecular Biologist

Bioinformatics

Note of Caution !!! Before we progress

You are entitled to study the following programs/tools/web server and its working methodology

NCBI – Entrez

GenBank

BLAST and its types

MapViewer

OMIM

dbSNP

And other available programs of NCBI

Molecular Biologist Point of View

KEY POINT

A phenotype is expressed by a Genotype

Patient developed new symptom

Disease

Collect tissue or cells representing a developmental stage

Isolate mRNA

Produce cDNA

Insert this cDNA into a suitable vector

Produce cDNA clones

Sequence these cDNA insert from either end

Expressed Sequence Tags (ESTs)

Finding a Disease Gene

ESTSingle pass, short 300–500 nucleotide sequences

cDNA clones

cDNA inserted into vector

cDNA

RTase

mRNA

RE

Sequencing

Finding a Disease Gene

Human Genome

Search

ESTs

Results

XYZ gene

XYZ gene

XYZ gene

KEY POINT

The XYZ gene expresses these ESTs.

Finding a Disease Gene

XYZ Normal geneT

XYZ gene variants

C

AG

T

SNPs

Normal

Genetics & Pharmacogenomics

To obtain information about the gene(s) causing the phenotype

Unknown EST collected from patient

Human Genome

Which BLAST to use ?

BLAST (human genome)

Genome (reference only) database Annotated Human Genome Assembly

MegaBlast

EST matches with a Contig

Query

What is a Contig ?

NCBI assembles component sequences from the human genome sequencing project into longer sequences called contigs whose accession numbers begin with prefix “NT_”

Annotated Human Genome Assembly

Component Sequences

Sequencing Projects

Assembly

Compare ESTs to The Human Genome

EST matches with a Contig

a real SNP or a sequencing error Position 16951392

Identify the Genes Expressing the ESTs“Known” genes annotated by alignment of EST and/or mRNA sequences to the assembly

The assembled genome contig sequence in the region

The Ab initio model genes predicted by the NCBI’s program Gnomon

The alignments of the known alternatively spliced transcripts

Genes_seq Map as a master map

Exons IntronsBLAST hit

HFE gene

Arrow downward=forward strand Arrow upward=reverse strand The HFE gene is annotated on the forward strand of chromosome 6

sv (Sequence Viewer), pr (Reference Proteins), dl (Download Sequence), ev (Evidence Viewer), mm (Model Maker), and hm (Homologene)

Variation Map as a master map

SNPs

Can you tell which SNPs corresponds to Exons and Introns ?Click any of the links and

obtain information about the location and the nucleotide variation

“Fasta sequence” and “Integrated maps” panels

SNP entry rs1800562

The location of the SNP, nucleotide position 16951392 on the contig NT_007592.14 of the reference assembly

Is the SNP non-synonymous ? GeneView Panel

The query EST sequence contains a known SNP in the HFE gene that results in a cysteine to tyrosine change in the 282nd amino acid

(Cys282Tyr) of the protein expressed by the longest HFE transcript variant

Gene

Alternatively Spliced Variants (mRNA)

SNPs

……..TAC…...

……..UGC……

Gene

mRNA

SNP

Tyrosine

Cysteine

Whether the HFE Gene Variant is Known to Cause a Disease Phenotype

The Cys282Tyr variant is reported to be associated with hemochromatosis

GeneSeeker

Cytogenetic Localization

Phenotype

Expression Patterns

Genes underlying human genetic disorders

List of Candidate Genes

GeneSeeker Methodology

DB Group-1 Localization dbs (Human)

GeneSeeker Methodology

DB Group-2 Localization dbs (Mouse)

GeneSeeker Methodology

DB Group-3 Phenotype & Expression Patterns

GeneSeeker Interface

GeneSeeker Result Page

Have a look at this Research Article

*Exhibition for Academic Studies

*

References

Identification of Disease Genes

Thank You For Your Attention !!!

It is freely available in HTML format

NCBI Bookshelf