Trans - Utility of Molecular Tools in Health Care and Biological Research Flashcards
discovered DNA double helix
James Watson, Francis Crick
BRCA 1 & 2
transcriptional molecules correlated with breast cancer (5% risk with BRCA 1; more risk with BRCA 2)
physical manifestation of genetic variation
anatomic variation
biochemical manifestation of genetic variation
physiologic
main genomic variations [3]
[1] single base pair change
[2] insertions, deletions
[3] structural rearrangements (ex. translocation)
epigenetic mechanism of genetic variation
[1] methylation of DNA
[2] histone packing
epigenetic mechanism affected by: [5]
[1] development (in utero, childhood) [2] environmental chemicals [3] drugs [4] aging [5] diet
methods of nucleic acid extraction
[1] organic-based extraction
[2] silica-based extraction
methods of nucleic acid amplification
[1] polymerase chain reaction
[2] cloning
post-amplification methods [4]
[1] direct assessment
[2] electrophoresis
[3] hybridization assay
[4] sequencing
when will you use organic methods of nucleic acid extraction?
when you want a greater quantity of nucleic acid
when will you use silica column method of nucleic acid extraction?
when you want greater purity of the sample
principle of nucleic acid extraction by organic solvent
[1] aqueous chloroform solution separates hydrophilic DNA from other components
[2] when alcohol is added, DNA precipitates at inferface –> easily collected
2 main organic solvents used in nucleic acid extraction
[1] chloroform
[2] alcohol
requirements of PCR [4]
[1] DNA template
[2] DNA polymerase
[3] DNA primers
[4] ions and pH buffers
PCR - temperature to activate DNA polymerase
94-96 C
PCR - denaturation step temperature
94-96C
PCR - annealing step - temperature and purpose
50-65 C to allow primers to bind to complementary strands
PCR - extension step - temperature and purpose
72 C to allow elongation
PCR - final extension step - temperature and purpose
70-74 C to ensure remaining single stranded DNA is fully extended
PCR - final hold - temperature and purpose
4-15 C for short-term storage
DNA polymerase used in PCR? why?
Taq polymerase - heat resistant polymerase from Thermus aquaticus
electrophoresis - principle
separation of DNA segments by size and charge via migration through gels driven by electrical charges
electrophoresis - purpose
to obtain a certain DNA segment of interest
electrophoresis - direction of flow? why?
towards the + terminal, because DNA is negatively charged
electrophoresis - rate of flow is affected by what factors?
rate of flow determined by size (larger = slower) and charge (more positive = slower)
LAMP - meaning
Loop Mediated Isothermal Amplification of DNA
LAMP - principle
PCR conducted at only one temperature
LAMP - principle
PCR conducted at only one temperature
LAMP - microorganism involved
bacillus stereos thermophiles (BST)
LAMP - microorganism involved
bacillus stereos thermophiles (BST)
LAMP - temperature
65 C
LAMP - temperature
65 C