What are STRs?

(short tandem repeats)

-usually 2-6 bp repeating DNA motifs

-highly polymorphic

Co-dominant inheritance

located in non-coding regions

How and why are STRs used in forensic science?

-High discriminatory power

-standardized loci sets

-works with small or degraded DNA

-Produces a numerical DNA profile for comparsion

What are the different classes/type of STRs?

-Simple: single repeated motifs

-compound: two adjacent motifs

-complex: repeats with internal vatiations

- by length: di-, tri-, tetra-

How does multiplex amplification work?

-Amplifies many STR loci at once

- Uses multiple primer pairs and different fluorescent dyes

-Requires careful primer design to avoid overlap/dimers

How are STRs used for identification?

-compare alleles across multiple loci

- A full profile has extremely low probability of being duplicated

-used to confirm or exclude contributors

- each person has a unique combination of STR alleles

-STR profiles are generated via capillary electrophoresis and compared to a reference sample or databases

-match probability is calculated using allele frquencies

Be able to calculate RMP given an electropherogram

-RMP product of genotype frequencies

-For heterozygotes: 2pq

-For homo: p^2

Multiply loci get overall RMP

Be able to describe how RMP is presented in court room

"The probability of two selected individuals having this profile is 1 in X"

- helps convey the strength of the DNA match to the jury

What are the different types of mixture analysis?

-qualitative: number of contributors, major/minor

-quantitative: estimating contributor ratios

-peak height analysis: relative contributor proportions

- statistical: likelihood ratios, probabilistic genotyping

Describe the different primer characteristics that contribute to PCR amplification?

-similar melting temps

-40-60% GC content

-high specificity

- Avoid hairpins, self dimers, cross-dimers

-short enough for degraded DNA
-length: 18-25 bg

How do you design a successful primer pair?

-Flanking target region, matching melting temperature, little complementarity, correct product size for multiplex

- Target conserved flanking regions

-Avoid repetitive or homologous sequencing

-Use software to optimise Tm, GC content, and specificity

- Validate with in silico PCR and empirical testing

What are the 2 types of DNA found in the cell? compare and contrast nuclear DNA and mtDNA

(nuclear DNA difference)

Nuclear DNA:

- inherited from both parents

- Location: nucleus

- highly individualizing

-linear

- diploid, low copy number

-used for STR profiling

What are the 2 types of DNA found in the cell? compare and contrast nuclear DNA and mtDNA

(mt DNA differences)

mtDNA:

- maternally inherited

-obtained from mitochondria

- circular shape

- high copy number-> survives degradation

- Not as discriminating

-used for degraded samples and maternal lineage

- more likely to have mutations

Why and when is mtDNA used in a forensic context?

- works when mtDNA is too degraded or absent

- Useful for hair shafts, bones, burned or old samples, maternal lineage tracing, missing persons

What is the HVI/II region?

-hypervariable regions of mtDNA control region

- most polymorphic parts-> best discrimination

-used for lineage tracing and degraded samples

How is mtDNA obtained?

1. collect sample

2. lyse open cells

3. DNA extraction

4. Amplification

5. Sequence the amplified mtDNA

6. compare to databases