Forensic Genetics in the 21st century – meeting the challenges of biological evidence

EUROFORGEN-NoE is funded by the European Commissionwithin the 7th Framework Programme

Peter M. SchneiderInstitute of Legal MedicineUniversity of Cologne, Germany

15/03/2015 Slide no 1

Forensic Genetics in the 21st century – meeting the

challenges of biological evidence


• The false idea that everything has been solved in f orensic DNA typing• can lead to miscarriages of justice due to confirmation bias – i.e.

correctly observed DNA profiles could result in wrongful conclusions • and may have contributed to the decrease in funding at national and

European levels

The 'CSI' Effect


The Growth of DNA Databases in Europe2003 – 2013

0

2.000.000

4.000.000

6.000.000

8.000.000

10.000.000

12.000.000

2003 2004 2005 2006 2007 2008 2009 2010 2011 2012 2013

Persons Stains

1.4 Millionunsolved crime cases

Persons +400%Stains +500%

Slide no 3


The European DNA Database Challenge

• How to catch unknown criminalswith known DNA profiles?

• Improve database workflow

• Harmonize legislation

• Enhance European data exchange

• Investigate new genetic tools

– to enhance resolution and sensitivity

– to predict biogeographic ancestry and externally visible characteristics

Slide no 4


HID-Ion AmpliSeq™ Identity Panel (A25643)

5

Identify degraded casework samples in less than 24 hr• Ready-to-use panel consists of 124 SNPs• Small amplicon size (average132;141 nt) for degraded DNA• Only 1 ng DNA recommended• 99.99% of concordance for 43 Ken Kidd SNPs

8 individuals per Ion 314™ chip




Concordance HID v2.2 – Inter-lab Comparison

98,2 98,2 97,895,3

98,2 98,5

1,2 1,2 1,54,1

1,2 1,5

75%

80%

85%

90%

95%

100%

Sample 1 Sample 2 Sample 3 Sample 4 Sample 5 Sample 6

6


Concordance HID v2.2 – Inter-run Comparison

97,63 97,79 98,2296,45

98,22 98,8297,04

1,78 2,21 1,783,55

1,78 1,182,96

75%

80%

85%

90%

95%

100%

NA06994 NA07000 HG00403 NA18498 NA07029 NA10540 NA11200

7


Concordance HID v2.2 – Sum of 'no-calls' and dropout s in sensitivity studies

8

Publication submitted and reviewed


Ancestry informative markers (AIMs)

• The EUROFORGENGlobal AIMs Panel

• 128 genetic markers• 5 major world populations• Suitable for forensic

casework– Sensitivity– Robustness– Speed

• Collaborative validation study using PGM™ system– Publication in preparation

Slide no 9


The Genetic "Photofit" Image

• Application of SNP panels for the prediction of externally visible physical traits, such as– Eye / skin / hair color– Hair morphology– Male baldness pattern– Body height– Facial characteristics

• More research required– Expensive studies– Large numbers of samples and analyses

needed to reach statistical power

Slide no 10

Modeling 3D facial shapes from DNA: Claes et al., PLOS Genet. 2014


Blood stains …?


Combined DNA/mRNA Work Flow

Identify and collect the biological sample

Co-extract DNA and RNA

DNA: PCR amplificationof STR markers

Fragment analysis byCapillary Electrophoresis

mRNA: remove residual DNA

with DNAse

cDNA: PCR amplification

of expressed genes

mRNA: reverse transcription

into cDNA


Complications when analysing mRNA

• mRNA levels vary: per cell type, per person, with physiological condition – Risk of missing cell type inference when using few markers � Several markers per cell type required!

• mRNA expression is enriched but not restricted– Risk for false positive results (if assay is overloaded)– Risk for false negative results

(if proportion of human RNA in assay is too small) � Replicates and clear interpretation guidelines required!


NFI: mRNA 20plex

• Blood – 3 markers• Saliva – 2 markers• Semen – 2 markers• Skin – 3 markers• Menstrual secretion – 3 markers• Vaginal mucosa – 3 markers• General mucosa – 1 marker• Housekeeping control genes – 3 markers�� 4 replicates of cDNA analysis

Blood Saliva Semen Skin

HBB CD93 AMICA1 STATH HTN3 SEMG1 PRM1 CDSN LOR LCE1C

amp 1

amp 2

amp 3

amp 4

Count 8/12 7/8 0/8 11/12

Menstrual secretion Vaginal mucosa Mucosa Housekeeping

MMP7 MMP10 MMP11 HBD1 MUC4 CYP KRT4 ACTB 18S GAPDH

amp 1

amp 2

amp 3

amp 4

Count 1/12 0/12 4/4 12/12

Positive detectionof body fluid onlyif > 50% of allmarkers and replicates are observed!

Specificity of mRNA detection


mRNA for special cases only?

• The scenario of a case or the type of sample has to be considered

• rape / sexual assault cases are most complex• Property crime typically does not involve intimate body fluids

– Large multiplexes not always necessary

• Small multiplexes with special applications may be easier to handle

• Skin/Saliva mRNA Pentaplex – Electropherogram of a skin (A) and saliva sample (B)


Skin/Saliva mRNA Pentaplex

Sample type LCE1C LOR CDSN HTN3 STATH

Skin-1 + - - - -

Skin-2 - - - - -

Skin-3 + + + - -

Skin-4 + + - - -

Skin-5 + + + - -

Skin-6 + + + - -

Skin-7 + + + - -

Skin-8 + + + - -

Skin-9 + + - - -

Skin-10 + - - - -

Saliva-1 - - - + +

Saliva-2 - - - + +

Saliva-3 - - - + +

Saliva-4 - - - + +

Saliva-5 - - - + +

Saliva-6 - - - + +

Saliva-7 - - - + +


Conclusions and Perspectives

• mRNA analysis provides a novel investigate tool for forensic casework– DNA: WHO has deposited a crime scene sample?– RNA: WHAT was the biological source of a sample?

• May help to reconstruct a crime event– But beware of association fallacies!

• Requires additional skills in handling and investig ating biological samples

• Application may be restricted to certain cases with challenging scenarios– Analytical strategy has to be decided prior to extraction procedure


Study on DNA profiling success rates

• To determine the relative chance of obtaining a DNA profile per sample category using data of six EUROFORGEN laboratories.

• A total of 27,401 casework samples analysed after December 2012 in six forensic laboratories is used.

• 44 categories of typical crime scene samples, each sample category containing data from 16 to 7,925 samples.

• Blood stains, cigarette ends and the collar of a coat are more useful sample types than the handle of a knife, a plastic bag or ‘plugs and cables’.

• All samples involving human body fluids or tissues generally contain higher amounts of DNA.

• 32% of analysed samples are contact traces, and about 20% produce informative partial/full profiles


DNA profiling success rates of typical crime scene samples

Murderers and rapists will attempt to have their convictions overturned after the Home Office admitted that potentially misleading DNA evidence had been presented to juries. The admission came five months after a leading forensic scientist alerted the government to a series of cases in which courts were given subjective summaries of complex DNA evidence rather than direct access to robust statistics.


DNA Evidence in the Courtroom

The Evidence Challenge

• A DNA profile was unambiguously assigned to a person

�� Sub-source level

• The DNA profile was recovered from a from a knife handle …

– has the DNA profile originated from skin cells?

�� Source level

• Were the skin cells deposited during the stabbing of the victim?

�� Activity level

• Were the cells deposited when the knife was used to cut a tomato?

• Were the cells deposited by secondary transfer?

• Was the DNA profile recovered from skin cells, or from a saliva trace?

Slide no 24


DNA Evidence in the Courtroom

The Association Fallacy• The strength of the evidence assigned to the origin of the DNA

profile recovered from the knife handle (sub-source level) is associated with the activity level (stabbing of victim) without considering alternative explanations

• Research urgently required on– DNA transfer (direct / indirect)

– DNA persistence (half-life of DNA profile)

– Body fluid / tissue source association with DNA pro file

Slide no 25


Thomson Reuters Science Watch: Forensics


First 20 institutions in citations and impact facto r: 14 European, 5 USA, 1 Australia


Percentage of papers published in Forensic Science Internationaland Forensic Science International: Genetics from 2009-2013

?

Forensic Publication Statistics


USA: Forensic DNA Research Funding (NIJ)

Slide no 29


The Virtual Institute of Researchin Forensic Genetics

Slide no 30

>180 Laboratories participating• Many potential research

partners …• Many potential research

ideas …

• Have you already joined?




Perspectives 2012-2016 and beyond

EUROFORGEN-NoE has initiated a wide array of activities• Networking (the virtual institute)• Original ground-breaking research

– Exemplar projects– Open call funded, but many more research ideas remaining unfunded

• Train-the trainer workshop series– Funding central training workshops– No funding for dissemination at national level

• Online resources for training and education– Online learning tools will require more money than available

• Ethical and legal aspects, and the societal dimensi on of forensic genetics

Slide no 33

EUROFORGEN-NoE is funded by the European Commissionwithin the 7th Framework Programme Slide no 34

Thank you for your attention!


Speaker was provided travel and hotel support by Thermo Fisher Scientific for this presentation, but no remuneration.

15/03/2015 Slide no 35

Life Technologies and its affiliates are not endorsing, recommending, or promoting any use or application of Life Technologies products presented by third parties during this seminar. Information and materials presented or provided by third parties are provided as-is and without warranty of any kind, including regarding intellectual property rights and reported results. Parties presenting images, text and material represent they have the rights to do so.

Ion PGMTM and associated HID panels are For Research, Forensic or Paternity Use Only. Not for use in diagnostic procedures.

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Forensic Genetics in the 21st century – meeting the challenges of biological evidence