The Omics Revolution Flashcards
List the classic measures of toxicity (6 measures)
- Histopathology
- Clinical Chemistry
- Metabolism
- Physiology
- Enzymology
- Electron Microscopy
List the new measures of toxicity (2 categories, 3 measures each)
- Gene expression:
- RNA Microarrays
- RT PCR
- Deep sequencing
- Protein expression:
- Western Blot
- Eletrophoresis
- ChIP
Define the goal of the various “Omics”
To determine whether the expression of genes, RNAs, proteins or metabolites (“signatures”) can serve as markers to predict toxicity
Define toxicogenomics and its three principle goals
The effects of a toxicant on all of the denes of a cell or tissue at the DNA, RNA protein, and metabolite levels.
- Understand the relationship between environmental stress and human disease susceptibility
- Identify useful biomarkers of disease and exposure to toxic substances
- Elucidate the molecular mechanisms of toxicity
Mow many of each histone are on a nucleosome, and how many base pairs does it coil?
2 of each (H2A, H2B, H3, H4) octameric
147 bp
What is the difference between Transcription and Translation in terms of gene expression?
Transcription: DNA -> preRNA
Translation: RNA -> protein
True/False? Gene expression precedes protein changes and toxicity
True
True/False? Changes in gene expression are not measurable at low doses
False
List the route of a typical toxicogenomic flow scheme (3 steps)
1) Exposure
2) In vivo study (dose/response time course)
3) Gene expression
- Amount of genes perturbed by candidate in each organ
- Signalling pathways representing significantly perturbed genes
- Compare with toxicogenomic database for predicted toxicological effects based on similarities of gene expression profiles
What are the 6 applications of a toxicogenomic flow scheme?
- Deciphering mechanism of action/pathway analysis of toxicant
- Response at low doses
- Revealing potentially novel health effects
- Identification of perturbed pathways -> targeted follow up
- Biomarker discovery
Predictive toxicogenomics
What year did affymetrix debut in?
1993
List the two types of microarrays and give examples of each.
Spotted chips:
- cDNA or PCR products
- Oligonucleotide
Perfect match/mismatch
Long oligonucleotide
Short oligonucleotide
On-Chip Synthesis:
- Photolithography (affymetrix)
What are the six advantages of microarrays?
- Eliminate questionable or low-quality measurements
- Remove probes in background
- Adjust the measurements to facilitate comparisons
- Select genes that are differentially expressed between samples
- Identify the biological processes and molecular functions that are alteredd
- Place data in the context of a health outcome
Define the p-value
P(data|H0 is true)
Probability of getting the same results givne that there is no link
low p-value incurs statistical significance
What are 4 reliable ways to determine significant changes in expression?
- Fold change
- t-test/ANOVA
- Permutation test (MANOVA or SAM)
Define GO
Gene Ontology, a controlled vocabulary of terms for describing gene product characteristics and gene product annotation data (includes cellular compartment biological function, and molecular process)
What does the KEGG pathway record?
everything
- metabolism
- genetic information processing
- environmental information processing
- cellular processes
- human diseases
- Drug development
- Toxicity assessment
What are the 10 steps in designing an experiment to study mechanism of action?
- Adequate sample size!
- Appropriate selection of time points
- Appropriate selection of treatment conditions
- Appropriate tissue/cells sampled
- Sample collection - randomization or block design
- High Quality RNA!
- Randomization and microarray experimental design
- implementation of QA/QC
- Appropriate normalization/filtering
- Validation with alternative technologies!
Possible reasons for discrepancies in published studies? (6)
- Lenient filtering methods for poor or low intensity spots
- Inclusion of saturated signals in analysis
- incorrect probe matching
- Improper data handling
- incorrect statistical analysis
- insufficient biological validation
Limitations of toxicogenomics? (6)
- Difficulty in analysis of high density data
- Difficulty in integration of data obtained by different technologies
- Difficulty in linking “omics” data to specific adverse effects
- Difficulty in translating statistical assessments into biological understanding
- Limitations of incomplete functional annotation of genome databases
Incomplete knowledge of functional pathways and networks, particularly trans-genome relationships
Methylation of DNA histones typically _______ gene expression + name the two enzymes responsable
Suppresses.
SAM - S-adenosylmethionine
DNMT - SNA methyltransferase
List the 5 histone modifications
Acetylation Methylation Ubiquitination Sumoylation Phosphorylation
5 types of non coding RNAs
miRNAs piRNAs siRNAs lncRNAs tsRNAs
