Download Document

Paola CASTAGNOLI
Maria FOTI
Microarrays. Applicazioni nella
genomica funzionale e nel
genotyping
DIPARTIMENTO DI BIOTECNOLOGIE E BIOSCIENZE
Microarrays: Applicazioni nella Genomica Funzionale e nel Genotyping
** ***
SAGE
Nylon membrane
Illumina
Bead Array
Different
Technologies
GeneChip Affymetrix
cDNA microarray
Agilent: Long oligo Ink Jet
CGH
Affymetrix Gene Chips
Whole genome
expression
Expression
variability
Whole
genome
genotyping
Sequence
variability
Mapping 10k, 100k and 500k
10,000, 100,000 and 500,000k
genotypes
Analysis of Genomics, Microarrays gene
expression and proteomics data contained in
public databases can identify the gene involved
in a particular human disease
2D gel
Only one candidate
gene
Disease
Gene Identified
with mutations
Computer
Search
Microarray
Gene expression data
Identification the candidate genes involved in
human diseases
More than one
candidate gene
NCBI
National Center for biotechnology
information
SNPs
Single Nucleotide
Polymorphism
Haplotype Blocks in Human
Genome
a)
a) Each human
chromosome is made
up of regions called
haplotype blocks 3 to
7 variants occuring at
a frequency of 5% in
the population. Each
haplotype block has
different colors
b) Two individuals with
differents haplotype
carrying 30% of
variation of the total
haplotype diversity in
the human population
Haplotype block
b)
Individual 1
Individual 2
Applications of Microarray
Technology
• Gene expression profiling
–
–
–
–
In different cells/tissues
During the course of development
Under different environmental or chemical stimuli
In disease state versus healthy
• Molecular diagnosis:
– Molecular classification of diseases
• Drug development
– Identification of new targets
• Pharmacogenomics
– Individualized medicine
Applications of microarrays
• Cancer research: Molecular characterization (taxonomy) of
tumors on a genomic scale
 more reliable diagnosis and effective treatment of cancer.
• Immunology - bacterial infections: study of host / pathogen
interactions in terms of their transcriptional response
• Developmental biology:
• Molecular biology: (conditional) gene knockout - find
regulated genes
• Response of cells / tissues to drugs / injury
Improvements in Medicine
• Diagnostics
–Faster and more accurate diagnosis
–Positive prognosis
–Predictable symptoms
–Ability to make arrangements
before things happen
Improvements in Medicine
• Disease Control and Prevention
–Increased knowledge about
genetic diseases
• Vaccines
• Treatments
• Education programs
Poor signature
Good signature
Gene Expression “Signature” as
a Predictor of Survival
70 reporter genes
Molecular Classification and Outcome
Prediction of Acute Leukemia in the
Genomics Era:
A Paradigm for Molecular Medicine
Cancer Molecular Medicine
Tailoring Therapy to the Disease and the Host
Genomics

Integration of Molecular, Genomic, and Ultimately Functional
Biologic Approaches into Diagnosis



Clinomics
Assessment of Genetics, Gene Expression Profiles for
New Molecular Classification Schemes and Identification
of Diagnostic and Therapeutic Targets
Assessment of Host Polymorphisms to Assess Disease
Risk and Potential Therapeutic Efficacy or Toxicity
Tailoring Therapy to Disease and Host
Uses of DNA Microarrays
• Transcriptome profiling:
– Massive parallel analysis can reveal patterns of gene expression
allowing researchers to predict gene associations and pathways
– Microarrays have been used over the past five years for transcriptome
analyses in organisms ranging from bacteria to fungi to humans to
higher plants under hundreds of different conditions
• Diagnostics: RNA or DNA samples hybridized to small arrays to
detect expression of marker genes (stress/pathogens,
developmental events)
• Molecular genetics: Genomic DNA samples are hybridized to a
limited number of probes to detect single nucleotide polymorphisms
(SNPs) for mutational analyses (pedigrees, gene mapping)
• Gene promoter analyses: ChIP to Chip assays – overlapping or
‘tiled’ DNA probes representing an entire genome are hybridized
with labeled, putative trans-acting proteins to detect binding to cis
elements in promoter regions