Topic 2 - Practical DNA Processing Flashcards
what 3 things must be done when a piece of evidence enters the lab
it must be recorded, photographed and sketched with annotations = ensuring a log of the item exists
note any damage
decide what presumptive tests you might want to perform
what is differential DNA extraction
an extraction method used when you are dealing with a mixed DNA sample that requires the separation of male and female DNA
what is laser microdissection and what does this allow us to do
a microscope with a laser attached allowing us to identify and cut out sperm heads
what is a DNA profile compared to initially
profiles currently on the National DNA Database
this contains profiles from those who have been arrested for recordable offences
when can two DNA profiles not be compared against one another
when different/previous methods of DNA profiling have been used
give 7 things done in DNA labs to prevent or identify contamination of samples
handle one sample at a time
wearing PPE - facemask, mob cap, gloves, lab coat, gloves
anyone working in the lab giving elimination samples
a separate room to change in and out of PPE
cleaning of workstations and equipment after use with a sample using alcohol
changing gloves
each bench has its own set of equipment that is logged so if contamination is found it can be traced back to an instrument
what is the name of the piece of equipment used for electrophoresis
a genetic analyser
why is preventing contamination more important as newer methods of DNA profiling are developed
newer methods are more sensitive so even the slightest contamination may be detected and can lead to misinterpretation of profiles
what was the phadebas test replaced with
the ELIZA test
give a piece of equipment that are a big source of contamination in labs
scissors
stains are not always obvious, so what can be used to after looking under ambient lighting to look for stains
alternative lighting e.g UV
semen will fluoresce
what is used in sketches to note different types of bodily fluid
different coloured pens - each lab has its own code
how can you confirm if a suspected bodily fluid is semen
examine under a microscope and look for the presence of sperm heads
what is organic extraction
using organic solvents to break down cells and isolate DNA
what is solid-phase extraction
using specialised columns e.g silica or magnetic beads to selectively bind to DNA and wash away the rest of the components
what three things does the choice of extraction method rely on
sample type
desired yield
what analyses are being used subsequently
when is differential extraction used
when a sample is likely to contain sperm
what is the extraction negative
a blank sample that is extracted using the same reagents and protocols as the samples
this should produce 0 peaks and suggest the extraction has been successful
what is the extraction negative used to identify
contamination sourced from the reagents or instruments or person doing the analysis
what is DNA quantification
how is this done
calculating how much DNA is present in a sample
done by adding a fluorescent probe or dye to the sample and comparing the fluorescence to a control sample or standards loaded into the equipment
what does the addition of fluorescent dyes allow us to measure
RFU’s - relative fluorescent units
name three types of quantifier often used in practice from thermofischer
quantifiler = measured human DNA
PicoGreen = not human specific but quick and easy
HY = indicates how much male DNA is present
what is the purpose of a positive and negative pressure lab
the pressure in the lab is higher than the corridor so hair will diffuse out not in
air can cause contamination by transferring DNA
what is the difference between denaturing and annealing
denaturing = splitting of strands
annealing = joining of two strands
what piece of equipment is used in PCR
a thermal cycler
what is a PCR positive
a sample with a known concentration of DNA
shows if the PCR has been successful and monitor performance of the thermal cycler
what is used to translate the peaks in an electropherogram to a series of numbered alleles
computer software
upon comparison with an allelic ladder also being run through the genetic analyser
what is an allelic ladder
a sample with known fragment lengths of DNA
was does ISS stand for
international size standards = specific DNA fragments of known sizes defined and used o size unknown fragments
