diagnostics 3 Flashcards
benefits of molecular diagnostics
provides info on origin of virus
helps ID new viruses and viral load
help design vaccines
determine transmission for zoonotic viruses
PCR
Polymerase Chain Reaction
makes multiple copies of gene segment
fast and inexpensive, amplifies DNA gene of interest
PCR steps
- Denature DNA, primer for both ends
- Taq Polymerase copies two new strands annealed to two old ones, making each complete strand semiconservative
- strands are elongated for 30-40 cycles, each cycle the amount of copies doubles
Real Time PCR
an advanced type of PCR which is quantitative, the target molecules are labeled with fluorescent dyes and can be counted, quick and easy like PCR
Genome Sequencing
Most common method is dideoxy Sanger
maps the genome of an organism
only way to tell between different species of viruses
can also ID genetic recombination, reassortment, and mutations present
Next Generation Sequencing
gradually replacing regular genome sequencing
do the same job with less DNA necessary, quicker, cheaper, more accurate, high throughput. Ex. Solexo
Metagenomics
study of the collective set of microbial populations in a sample by analyzing the samples entire nucleotide sequence content. Usually next generation sequencing is used and this can help ID new pathogens
Why is genome sequencing important?
pathogen detection, studies on genetic variation, ID novel strains, development of diagnostics, ID gene associated with drug resistance, development of therapeutics, show strength of vaccines
phylogenetic analysis
use of virus genomes to study evolution of viruses and possible genetic relationships
ex. porcine rotavirus genetically close to human rotavirus
microarray analysis
a way to probe for hundreds of pathogens using one test where fluorescent labeled targets are annealed to DNA fragments on the silicon tray and color indicates positive annealing, or presence of that pathogen
