Complex STR profile interpretation

Question 1

Name the 4 ways mixtures can occur within a sample

Answer 1

1. Victim and suspect DNA on a single sample - sexual assault
2. Multiple suspect DNA on a single sample - gang rape
3. Suspect and background DNA on a single sample - victims DNA from a robbery
4. Contamination of sample during handling

Question 2

Name 5 procedures to avoid mixture analysis

Answer 2

1. Differential extraction during DNA recovery
2. Laser microdissection
3. Use of quantification kit with mixture detection capabilities
4. Y-STR profiling
5. Processing of additional non-mixed stains

Question 3

How do we determine if a STR profile contains a mixture

Answer 3

• more than two peaks at a locus
• severe peak imbalance between heterozygous alleles at a locus
• abnormally high stutter

Question 4

How do we asses the extent of the mixture after identification

Answer 4

by using the quantitative peak height information that is provided by the capillary electrophoresis

Question 5

From a simple two person mixture, what are the two kinds of major/minor contributors

Answer 5

1. Major/minor
2. Equal major

Question 5

Within a simple two person mixture what are the two allele contributions described as and why

Answer 5

1. Major component - larger quantity of that allele in the mixture
2. Minor component- smaller quantity of that alleles makes up the mixture

Question 6

How do you calculate the mixture ratio

Answer 6

sum of peak heights of major/ sum of peak heights from minor

Question 7

what does the mixture ratio of an STR profile describe

Answer 7

the extent of the different amount of each alleles in the mixture

Question 8

How do you calculate the mixture proportion of an STR profile

Answer 8

(sum of peak heights major/ sum of all peak heights) x100

Question 9

If one allele is observed at a locus, what 2 way can it be interpreted, and how do we distinguish between them

Answer 9

1. single homozygote
2. two homozygotes with overlapping alleles
   - need to look at the full profile to asses which is more likely E.g. good heterozygote balance throughout the STR profile would suggest a true homozygote

Question 10

If two alleles are observed at one loci, what 4 ways can this be interpreted

Answer 10

1. Single heterozygote (no mixture)
2. Two heterozygotes with overlapping alleles
3. Two homozygotes with non-overlapping alleles
4. Heterozygote and homozygote with one overlapping allele

Question 11

What are the two main stochastic effects at low level (DNA artefacts)

Answer 11

1. allelic drop-out - losing an allele
2. allele drop-in - presence of an additional allele

Question 12

What is the mechanism behind allelic drop-out

Answer 12

the random collection of template material during pipetting and the random preferential amplification of an allele during PCR

Question 13

What is the mechanism behind allelic drop-in

Answer 13

airborne material entering the mix before PCR or airborne PCR product entering the reaction before CE

Question 14

What causes allelic drop-in/out

Answer 14

uncontrollable events - E.g. the chance of DNA polymerase interacting with an allele

Question 15

How do we best manage the uncontrollable events that can cause allelic drop-in/out

Answer 15

repeat runs of the same DNA sample and report consensus sequence

Question 16

What is a null allele

Answer 16

an allele that is present within a sample but is not amplified due to a primer-binding site mutation - therefore the allele will not appear in the STR profile

Question 17

What does an ‘off-ladder’ allele refer to

Answer 17

an allele that does not conform to a bin window - they are between a whole STR repeat unit

Question 18

What can cause off-ladder alleles

Answer 18

the presence of a recognised partial repeat due to mutation

Question 19

What are the two types of tri-allelic patterns

Answer 19

1. One allele is roughly double the other two
2. All three alleles are roughly equal

Question 20

What do inheritance studies suggest about tri-allelic patterns

Answer 20

1. They occur due to the inheritance of two chromosomal regions from one parent
2. They can be observed in paternity analysis

Question 21

What are dye artefacts and how do we distinguish them in an STR profile

Answer 21

peaks that are not caused by PCR product but from instrumentation
- Usually they display a different morphology to an allele - peak is not normally distributed
- They do not have stutter peaks

Question 22

What does mixture analysis require

Answer 22

expert software