FSCI6603 - Analysis of Mixtures

7/28/2019 FSCI6603 - Analysis of Mixtures

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FSCI6603


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Mixtures arise w en more t an one in ivi uacontributes to the DNA stain

Interpretation of forensic DNA mixtures is a

challenging task in forensic casework

Laboratories should conduct sufficient

-

.


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Common in cases of sexual assault where thesource of DNA evidence can include: e v c m,

the perpetrator(s), and

the consensual artner(s) of the victim

Even more complicated in incest cases

http://www.nfstc.org/pdi/Subject06/images/pdi_s06_m03_06.swf


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It is also possible to find a mixture in aknown blood sample from a deceased,

to death

It is also possible to find a mixture whenintimate sam les are anal zed Intimate samples are generally swabs collected

from a persons body (skin, buccal, vaginal swabs)


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possible allele overlap among an unknownnumber of contributors

-a e e patterns

stochastic fluctuation with low quantity or

the semi-quantitative activity of Taqol merase


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A stain exhibiting more than two alleles in a

single DNA system shall be considered amixed stain except in the case of genetic. ., ,

mosaicism, or duplication)

If more than two alleles are observed in atleast two DNA systems, the presence of a

mixed stain shall be assumed


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When there are three or more alleles areobserved at multiple loci

Notable differences in the allele intensities


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The presence of more than two alleles at anyone locus

pea n a stutter pos t on t at as a g erRFU than what would be expected, based

that locus (>50 RFU)

Imbalanced and/or unex ected eak hei ht

percentages


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No obvious major

evidence of stochasticeffects

Class A Stochastic effects


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Clearly distinguishable Mixtures with no major

components;consistent peak height

evidence of stochasticeffects

ratios of approximately4:1 (major to minor

heterozygous systems,and no evidence of

Class B Class C


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ocus represent aclear major and minorcontributor

(vWA locus) representan am guous m norcontributor

vWA major contributormas ng m nor con r u or


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STR Typing Results of the Forensic Evidence (Sperm Stain) andthe Reference Samples from the Victim and Suspect*.

Locus Victim Semen Stain SuspectD3S1358 15, 19 15, 17, 19 15, 179 9 3, . 9, 9.3 , .D21S11 31, 32.2 29, 30, 31, 32.2 29, 30D18S51 12, 16 12, 16, 18 12, 18Penta E 11, 14 11, 14, 17 14, 1711, 11, ,D13S317 11, 11 11, 13 11, 13D7S820 11, 12 9, 11, 12 9, 11D16S539 11 11 10, 11 10 11,CSF1P0 12, 13 11, 12, 13 11, 13Penta D 11, 12 11, 12 11, 12VWA 17, 18 15, 17, 18 15, 17

10 13, 10, 13 ,TPOX 8, 8 8, 8 8, 8FGA 21, 23 21, 23 23, 23Amelogenin X, X X, Y X, Y*Alleles given in bold could be assigned to the suspect upon re-analysis


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In general, the presence of not more than foura e es n a g ven system a ows t e assumpt onof at least two independent stain donors

In general, the presence of not more than six

alleles in a given system allows the assumptionof at least three independent stain donors

,in a given system, the exact number of staindonors cannot be reliably determined


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No universal formula used to interpret andreport mixtures

Once the determination of a mixture has,information and/or sample type to aid in theinter retation of the sources of the mixture


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It is common to obtain samples where one ofthe contributors (e.g., the victim) is known. In,

unknown profile by subtracting the

mixed profile

http://www.nfstc.org/pdi/Subject06/images/pdi_s06_m03_06_b.swf


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Allele Peak Height19* 250

20 674

23 717

25* 225


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Assuming only two donors, the mixtureproportions are estimated by summing thepeak heights of the two major or minor

four alleles. This is then multiplied by 100 to

rovide a ercenta e contribution. 250 + 225 / 250 + 674 + 717 + 225 = .25 x 100

= 25% contribution from the minor donor

= .= 75% contribution from the major donor


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This can also be represented as a ratio bysumming the peak heights of the two minor

peak heights from the two major alleles

250 + 225 / 674 + 717 = .34 or a ratio ofapproximately 1 to 3


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A second check of this hypothesis isevaluation of the peak height percentages. Inthis case the peak height percentage for

,(20, 23) is 94%

These values are within the acceptable rangeof peak height percentage for balanced

heterozygotes


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If the genotypes of two persons are abandbc, then they share ballele. Thecontributions are assumed to be additive

Can expect the proportions ofa:b:c= 2:3:1


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pp y ng stat st ca nterpretat on s s mp e w en

dealing with one contributor (e.g. single sourcesamples or mixtures where a major componentcan be defined or inferred).

more complex

e as s or a ca cu at ons s t e now e ge othe allele frequencies in the relevant population


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Perform no statistical calculations : Thesuspect is included or excluded from being a

Performing match probability estimations

contributors

Usin exclusion or inclusion robabilities (PE)

or (PI)

Performing likelihood ratio calculations (LR)


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DNA stains from crime scenes are usuallycharacterized through multi-allelic (STR) loci,

approach is the most efficient in determining

mixture


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If all alleles of a person If alleles of a person in

uniformly present in amixed stain, the

present in a mixedstain, the person shall

person shall beconsidered a possible

not be considered as apossible contributor to

Inclusion Exclusion


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PI represents the combined probability(relative population frequency) of allcombinations of genotypes that cannot be

profile of a stain

PI is equivalent to the match probability in thecase of a stain originating from a single

person


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It is equivalent to the probability that arandomly selected person is a contributor to

[=random man not excluded (RMNE)].


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PI is independent of assumptions about thenumber of possible contributors to a stain,

,persons involved in a given case

Based on the criteria given for inclusion


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The probability of exclusion provides anestimate of the proportion of the populationthat has a genotype composed of at least one

=

indicates the probability of excluding arandomly selected person as a contributor to

a given stain


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PI is calculated from the sum of all genotypes of

Assumin that allele fre uenc data conform toHardyWeinberg equilibrium

Assuming a frequency of 0.1 for all alleles

PI 0.32 0.09 or 9%


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Expected that 9% of a group of randomlyselected persons will not be excluded as stain

~person (RMNE)

The PE is calculated from the difference

PE=1

PI = 1- 0.09 = 0.91 or 91%


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Expected that 91% of a group of randomlyselected persons will be excluded as stain


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The likelihood ratio is the ratio of twoprobabilities of the same event underdifferent hypotheses

Thus for events A and B, the probability of A

,probability of event A given that B is false (H2)gives a likelihood ratio


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Based on the assumption of two mutuallyexcluding hypotheses

Requires the description of a distinct scenario

Both hypotheses explicitly describe


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Likelihood ratios are usually constructed withthe numerator being the probability of theevidence (E) if the identified person is the

,being probability of the evidence (E) if an

unidentified erson is the source of theevidence

LR = L (EIH1)L (EIH2)


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P(E/H1

) is the probability of the evidencegiven a presumed individual is the

2 given the presumed individual is not thecontributor of the evidence


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Given a two-person mixed stain M and thata o serve a e es can e exp a ne y e

genotype of the victim, Gv, and the genotypeof the sus ect Gs the h otheses can beformulated as follows:

M originates from the victim Vand the suspect S

originates from the victim Vand from a unknownperson Uunrelated to the suspect


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LR = L (MIH

p) = L (MIGv,Gs)L (MIHd) L (MIGv, Gu)

The resulting LR provides a numerical value,which indicates how many times more likelythe observed DNA profile is under the

compared to the scenario described in Hd


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First derive the numerator of the LR.

The prosecution claims that the stain can beexp a ne y a com nat on o t e genotypesof the victim and the suspect, as there are no

Hence the numerator results as

L(MIHp) = L(MIGv,Gs) = 1


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The defense, however, claims that thesuspect has not contributed to the stain.

The genotype of the suspect is not relevant

must be explained by the contribution of an

As allele c may have been contributed eitherby a person homozygous for allele c or from

a person heterozygous for c in combinationwith allele a or b,


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the denominator is as follows:

L(MIHd) = L(MIGv,Gu) =2ac + 2bc + c2

LR = 1

2ac+2ab+ + c2

Assuming the freq = 0.1 for all alleles

LR = 1 10.02+0.02+0.01 = 0.05 = 20


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it is 20 X more likely to observe the DNAprofile if the mixed stain originated from the

from the victim and an unknown person (or a


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LR < 1 the genetic evidence has moresupport from the denominator hypothesis

= t e genet c ev ence as equasupport from both numerator and

LR > 1 the genetic evidence has moresu ort from the numerator h othesis


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The likelihood ratio is a ratio of probabilities,and can take a value between zero and

,the first hypothesis (Hp) is true


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Establish mixture interpretation guidelines tominimize potential bias that could be

include mixture studies prior to performingcase work anal sis usin a new technolo


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Based on the evaluation of the profile in itsentirety

Distinguish true alleles from non-allelic

Stutter peaks

Minus A -A eaks Pull-up peaks

Off ladder alleles

ea e g m a ance


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The RFU can be used to establish peak heightand/or peak area

The RFU can therefore be used the determine

values for fluorescent signals


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Peak Amplitude Threshold (PAT)

Defines the minimum peak height in RFU thatcon ent y ascr es a true amp con peaand when confidence is too low to reliably

PAT the lowest peak height value that the labuses

PAT above the lowest limit of detection (LOD)


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Match Interpretation Threshold (MIT)

MIT establishes the minimum peak height int at a amp con pea must sp ay to

confidently conclude that no genetic

detected after PCR due to low template conc,degraded sample or inhibitors


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The PCR process is not 100% efficient

Non-allelic peaks such as stutter, pull-upscan be introduced into the process

For an unbiased assessment of the potential

the evidentiary profile(s) before working onthe reference sample(s)


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Allelic vs non-allelic determinations for the

evidentiary profile will therefore not be

bias predicated on the DNA profile of the


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A t oug uncommon can e o serve at a

loci in a profile and can be from a single

Most likely interpreted as a mixture?

Things to consider peak height imbalance

Must be considered on a case by case basis


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Estimate the minimum number ofcontributors

ase on t e oc t at ex ts t e t egreatest number of allelic peaks

E.g. 5 alleles are detected at 1 or more loci,the DNA t in results are consistent with

having arisen from 3 or more individuals


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The STR typing result for exhibit Q1 is a

mixture of DNA from three or more

the presence of DNA from at least threeindividuals


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Fingernail debris (recovered as scrapings) was taken from a manaccused of sexuall assaultin his biolo ical dau hter b di itallpenetrating her vagina.


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Fingernail scrapings VWA THO1 F13

16,18 7,9.3 5,7

VWA THO1 F13

Suspect 16,18 7,9.3 5,7

aug ter , . , . ,


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Ladd et al Interpretation of Complex Forensicx ures : - ,

P. Gill et al DNA commission of the

Recommendations on the interpretation ofmixtures. Forensic Science International 160

Budowle et al Mixture interpretation: Defining

the relevant features for uidelines for the

assessment of mixed DNA profiles in forensiccase work. J Forensic Sci 54: 810 821, 2009

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FSCI6603 - Analysis of Mixtures