Identification of Disease Genes Identification of Disease Genes Pharmacogenomics & Drug Design S.Prasanth Kumar, S.Prasanth Kumar, Bioinformatician Bioinformatician S.Prasanth Kumar Dept. of Bioinformatics Applied Botany Centre (ABC) Gujarat University, Ahmedabad, INDIA www.facebook.com/Prasanth Sivakumar FOLLOW ME ON ACCESS MY RESOURCES IN SLIDESHARE prasanthperceptron CONTACT ME prasanthbioinformatics@gmail. com
This document is posted to help you gain knowledge. Please leave a comment to let me know what you think about it! Share it to your friends and learn new things together.
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