Topic 2 - Practical DNA Processing

Question 1

what 3 things must be done when a piece of evidence enters the lab

Answer 1

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

Question 2

what is differential DNA extraction

Answer 2

an extraction method used when you are dealing with a mixed DNA sample that requires the separation of male and female DNA

Question 3

what is laser microdissection and what does this allow us to do

Answer 3

a microscope with a laser attached allowing us to identify and cut out sperm heads

Question 4

what is a DNA profile compared to initially

Answer 4

profiles currently on the National DNA Database

this contains profiles from those who have been arrested for recordable offences

Question 5

when can two DNA profiles not be compared against one another

Answer 5

when different/previous methods of DNA profiling have been used

Question 6

give 7 things done in DNA labs to prevent or identify contamination of samples

Answer 6

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

Question 7

what is the name of the piece of equipment used for electrophoresis

Answer 7

a genetic analyser

Question 8

why is preventing contamination more important as newer methods of DNA profiling are developed

Answer 8

newer methods are more sensitive so even the slightest contamination may be detected and can lead to misinterpretation of profiles

Question 9

what was the phadebas test replaced with

Answer 9

the ELIZA test

Question 9

give a piece of equipment that are a big source of contamination in labs

Answer 9

scissors

Question 10

stains are not always obvious, so what can be used to after looking under ambient lighting to look for stains

Answer 10

alternative lighting e.g UV

semen will fluoresce

Question 10

what is used in sketches to note different types of bodily fluid

Answer 10

different coloured pens - each lab has its own code

Question 10

how can you confirm if a suspected bodily fluid is semen

Answer 10

examine under a microscope and look for the presence of sperm heads

Question 11

what is organic extraction

Answer 11

using organic solvents to break down cells and isolate DNA

Question 12

what is solid-phase extraction

Answer 12

using specialised columns e.g silica or magnetic beads to selectively bind to DNA and wash away the rest of the components

Question 13

what three things does the choice of extraction method rely on

Answer 13

sample type
desired yield
what analyses are being used subsequently

Question 14

when is differential extraction used

Answer 14

when a sample is likely to contain sperm

Question 15

what is the extraction negative

Answer 15

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

Question 16

what is the extraction negative used to identify

Answer 16

contamination sourced from the reagents or instruments or person doing the analysis

Question 17

what is DNA quantification

how is this done

Answer 17

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

Question 18

what does the addition of fluorescent dyes allow us to measure

Answer 18

RFU’s - relative fluorescent units

Question 19

name three types of quantifier often used in practice from thermofischer

Answer 19

quantifiler = measured human DNA
PicoGreen = not human specific but quick and easy
HY = indicates how much male DNA is present

Question 20

what is the purpose of a positive and negative pressure lab

Answer 20

the pressure in the lab is higher than the corridor so hair will diffuse out not in

air can cause contamination by transferring DNA

Question 21

what is the difference between denaturing and annealing

Answer 21

denaturing = splitting of strands

annealing = joining of two strands

Question 22

what piece of equipment is used in PCR

Answer 22

a thermal cycler

Question 23

what is a PCR positive

Answer 23

a sample with a known concentration of DNA

shows if the PCR has been successful and monitor performance of the thermal cycler

Question 24

what is used to translate the peaks in an electropherogram to a series of numbered alleles

Answer 24

computer software

upon comparison with an allelic ladder also being run through the genetic analyser

Question 25

what is an allelic ladder

Answer 25

a sample with known fragment lengths of DNA

Question 26

was does ISS stand for

Answer 26

international size standards = specific DNA fragments of known sizes defined and used o size unknown fragments