Molecular Diagnosis Flashcards
What is PCR?
polymerase chain reaction
-quick and accurate technique to make numerous (millions to billions) copies of a piece of DNA of interest
What is quantitative PCR?
real-time PCR
-monitoring the amplification process
-helps detect expression level of a gene
What are the detection methods used in qPCR?
non-specific fluorescent dyes for double-stranded DNA
-CYBR Green 1
fluorescent reporter probe (more $$$)
-TaqMan Probe
What are the two types of DNA sequencing?
whole genome sequencing (WGS)
-coding and non-coding sequences
-cost: $1,000-$3,000
whole genome exon sequencing (WGES)
-coding sequences only
-cost: $1,000-$2,000
What kind of information can you get about diseases from DNA sequencing?
point mutations
deletions
insertions
SNPs
structural variations
detect family disease history
What is RNA sequencing?
study effects of transcriptome on phenotypes such as disease susceptibility
-cost: $500
What are the applications of RNA sequencing?
transcript identification
splice variant analysis
differential expression
clinical diagnosis
Describe the general schematic process for RNA sequencing.
extract RNA
make cDNA & shatter into fragments
map reads (puzzle together)
What is the largest gene in the body?
DMD
What is a biochip?
an array of selected biomolecules immobilized on a surface
-DNA or protein
-dont have RNA chips (unstable)
What is a microarray?
a rapid method of sequencing and analyzing genes
-DNA chip
What are the applications of biochips and microarrays?
DNA sequencing
drug discovery and development
molecular diagnosis
biomarker identification
drug safety
pharmacogenomics
What are some biochip technologies?
PCR on a chip
-customized microarray with miniaturized PCR reactor for amplification
gene profiling array
-human genotyping, CNVs, transcriptome analysis
Arrayit H25K
AmpliChip CYP 450
What is the NanoChip BRCA?
an array for diagnosis of risk for breast cancer
-14 most common mutations in BRCA1 and BRCA2 genes
What is 23andme?
global screening array that analyzes over 600,000 SNPs
-fixed markers: ~654, 027
-custom marker: up to 50, 000
provides information on health and ancestry
What is the ProteinChip?
first complete tool for disease-focused protein analysis
Which technology are protein chips based on?
SELDI (surface-enhanced laser desorption/ionization)
-surface enhanced selective capture
-high sensitivity time of flight mass spectrometry (TOFMS)
What size peptides and proteins can protein chips detect?
1,000-300kd
What is Tandem-MS commonly used for?
proteomics
What is microfluidics?
one of the most important innovations of biochip technology
-technology to precise the chip
based on advances of microfabrication
reduction in size and increase in throughput in sample analysis
What is the benefit of microfluidics?
increased reaction rate, enhanced detection sensitivity, and control of adverse effects
What is FISH?
fluorescence in situ hybridization
-use fluorescent probes to detect and localize a specific DNA sequence
-probe has a degree of sequence complementarity with the target
What is SNP genotyping?
high resolution genome-wide association of SNPs to risk profiles of common diseases
-SNP panels: 300,000-1,000,000
What are the associated studies with SNP genotyping?
SNPs and disease susceptibility
SNPs and drug responses
SNPs and treatment outcomes
How are CNVs detected?
high-throughput scanning technologies such as comparative genomic hybridization (CGH) and high-density SNP microarrays
What is gene expression profiling?
measurement of the expression and activity of thousands of genes at once (genomics and transcriptomics)
-get a global picture of cellular function
What can be identified with gene expression profiling?
association of gene expression profiles with disease susceptibility and development, drug metabolism and adverse effects
identify drug design targets and predict drug responses
What are examples of molecular imaging?
optical imaging (fluorescence, bioluminescence)
MRI scan
near-infrared imaging
single photon emission computed tomography (SPECT)
PET scan
What is a PET scan?
positron emission tomography
-positron: charge +1, spin 1/2
-use a radioactive drug as a tracer, such as fluorodeoxyglucose (glucose analog)
What are PET scans widely used for?
cancer diagnosis and treatment
