Reading...
Play button
Play button
Use LEFT and RIGHT arrow keys to navigate between flashcards;
Use UP and DOWN arrow keys to flip the card;
H to show hint;
A reads text to speech;
18 Cards in this Set
- Front
- Back
- 3rd side (hint)
|
What are the 3 steps in putting together a new molecular diagnostic test?
|
-choose target gene
-choose method of amplification -choose detection method |
|
|
|
What are the criteria for selecting a gene target?
|
-DNA/RNA/protein
-short (shorter amplification time) -60% GC content (stable interaction with probe) -highly conserved gene (must be found in organism) -gene-specific locus (must be specific to organism) |
|
|
|
Which is better for quantification, PCR or RT-PCR? How is RT-PCR different from PCR?
|
RT-PCR (for every cycle, 1 molecule will be detected by laser detection)
|
-probe has reporter and Quencher molecule that emit signal when separated
-when Taqman pol (5'-5' exonuclease activity too) comes to probe, it will cleave off the reporter, emitting fluorescence for the one strand it just amplified |
|
|
Describe real-time immuno-PCR
|
-detects protein
-antibody on plate, then add protein, then add second antibody with biotin -add Si which binds very strongly to biotin -then add DNA with biotin attached to bind to antibody-biotin-Si complex -run PCR |
|
|
|
What are the standards for determining whether or not a molecular diagnostic test is good or bad?
|
-accuracy vs. precision
-specificity vs. sensitivity |
|
|
|
Differentiate between sensitivity and specificity
|
-sensitivity: ability to detect people who actually have the disease
|
-specificity: ability to detect people who do not have the disease
|
|
|
What is the calculation for sensitivity? Specificity?
|
sensitivity = true positives/(true positives+false negatives) x 100
Sensitivity is indirectly proportional to # of false negatives |
specificity = true negatives/(true negatives+false positives) x 100
Specificity is indirectly proportional to false positives |
|
|
What is an ROC curve? What does it represent and what are the labels of axes?
|
-Receiver-operator characteristic curve
-represents trade-off between sensitivity and specificity, how good the diagnostic test is -further up in left corner the better |
-y axis = sensitivity (1-false negative rate)
-x axis = 1-specificity (false positive rate |
|
|
T or F: specificity is strongly influenced by prevalence of disease population
|
TRUE
|
|
|
|
What is a linear dynamic ranger?
|
linear range that can be used for accurate quantification
|
|
|
|
how can we test individual identity?
|
-short tandem repeat (STR)
-no individual shares the exact same molecular profile with others |
|
|
|
what are short tandem repeats?
|
-number of repeats at telomeres
-no 2 people have identical number so can be used to identify person of a crime |
|
|
|
what is the problem with STR testing?
|
very sensitive so contamination is likely
|
|
|
|
What is a melting curve? How can mutations be detected using a melting curve?
|
-melting curve plots % dsDNA against temperature
-melting point is where dsDNA=ssDNA |
-every DNA sequence has characteristic melting curve and Tm due to GC content
-with every mutation, the Tm changes |
|
|
Why is it beneficial to study a single cell rather than a section of tissue?
|
-tissue is heterogenous, don't know how much/where in tissue has tumor
-single cell is just one cell so simpler |
|
|
|
How can a single molecule (eg. DNA) be detected?
|
nanoparticle-conjugated fluorophores (bio-barcode assay, BCA)
|
|
|
|
How does bio-barcode assay (BCA) work?
|
-have magnetic particle that has DNA/RNA/protein of interest
-add gold particle with barcode DNA -capture with magnetic field -run through scanner to find molecule of interest -highest detection known today (zeptomolar) |
|
|
|
how can molecular based testing be used for personalized medicine?
|
-pharmacogenetics (unique prescription for each person)
-theragnostics (determines whether or not a drug should be used for certain therapy) -prognomics (patient prognosis for disease and after treatment |
|
