More on the Practical Aspects of DNA Profiling Flashcards
What is the unzipping enzyme in DNA replication?
- helicase
What enzyme are DNA fragments sealed together by?
- ligase
What are three tests used to generated DNA profiles?
- RFLP (restriction fragment length polymorphisms)
- DG alpha
- automated STR’s
What is RFLP?
- restriction fragment length polymorphisms:
- first DNA testing used (1980/1990s)
- produces auto-radiograph (x-ray plate and looked like barcode pattern)
- used radiation to mark where DNA bands are
- took a month to generate profiles
- very discriminating
What is HLA DQ alpha?
- first PCR-based test
- also called poly-markers
- critical information comes from a series of test strips
- look for presence of absence of blue dots (pattern of dots = what DNA molecules are present)
What are the benefits of DQ alpha over RFLP?
Is there any negatives?
- DQ alpha more sensitive in terms of starting material that is needed to generate a profile
- DQ alpha is faster (half a day compared to months)
- Yes DQ alpha had a lower discriminating power, RFLP was better for this
What is MLP?
What are limitations of this
- multi-locus probe (1985)
- lengthy
- required a large amount of sample to produce a profile
- results were complex to interpret
What is SLP?
What are advantages of this?
What are the limitations of this?
How are results interpreted if got four children and father?
- single locus probes (1987)
- easier to interpret
- initial results obtained in a few days but took over a week to prepare a full profile
- lacked sensitivity
- poor results with degraded samples
- difficult to resolve mixtures
- lane 3 - shares one band with each of the other lanes
Define sensitivity and discriminating power?
sensitivity = amount of starting material we can begin working with
discriminating power = power to distinguish between two samples
What is six criteria used for judging the benefits of a DNA profiling system?
- discriminating power
- sensitivity
- ability to deal with artefacts
- speed
- ability to deal with mixtures
- ability to conduct database searches
What is mitochondrial DNA?
What are the drawbacks of it?
What can be said about tests for mitochondrial DNA?
- it is more sensitive than automated STRs
- each cell has thousands of mitochondria associated with it
drawbacks:
- it is relatively small compared to the amount of DNA that is present within the nucleus and so not as much information is contained within
- it is maternally inherited (from mother)
- tests are very sensitive but not as discriminating because we know where we find one kind of mitochondrial DNA there will be many more maternal relatives who will share this genetic material
What is Y-STR?
What is a drawback?
- tests on Y-STR markers which reside on human Y chromosome (female XX and male XY)
- there is information on the Y chromosome that can be analysed by forensic scientists where there is possibility of male contributor to sample
- can give us insight into male DNA profile without seeing contributors from female
- helpful when talking about a mixed sample
- father son grandfather and great grandfather have same Y-STR profile
How much DNA do we leave behind in fingerprint?
How much DNA is transferred when wearing a garment?
How much DNA is needed for DNA17?
- 100 cells
- enough to generate a DNA profile using the widely available commercial DNA assays
- 1000 cells (at 6 pg per cell this is 5000 pg)
- 80 cells (500 pg)
- DNA profiles have been obtained from as few as 15 cells when these are in good condition
What are polymorphisms?
- non-coding regions of DNA which differ greatly between person to person
- many areas of DNA genome is same for everyone (no use for distinguishing between people)
- DNA in this region is likely to come in many forms and sizes
Define locus and allele
locus = a specific location on a chromosome
allele = a specific region of DNA which varies between people
- at each locus we see two alleles (if heterozygous) for an individual
