WILL NGS AND CE TECHNOLOGY ENHANCEMENTS

LEAD TO A SIGNIFICANT EXPANSION OF THE CORE LOCI?

Identity Benefits and Privacy/Policy /Legal Issues

Presented By:

Tim Schellberg & Bruce Budowle

ISFG – Krakow, Poland

August 31, 2015

These countries have implemented legislation/polices on a national basis to database the DNA of a defined category of criminal offender

AustraliaAustriaBahrainBarbadosBelarusBelgiumBrazilCanadaCzech RepublicChileChinaCroatiaCyprusDenmark

EstoniaFinlandFranceGermanyHong KongHungaryIcelandIsraelJapanJordanKuwaitLatviaLithuaniaNetherlands

New ZealandMacedoniaMalaysiaMauritiusNorwayOmanPanamaPolandPortugalQatarRussiaSloveniaSlovakiaSingapore

South KoreaSpainSwedenSwitzerlandTaiwanUnited Arab EmiratesUnited KingdomUnited StatesUruguay

51 COUNTRIES HAVE IMPLEMENTED NATIONAL PROGRAMS

OVER 60 MILLION OFFENDER SAMPLES

NEXT COUNTRIES TO IMPLEMENT NATIONAL CRIMINAL OFFENDER DATABASE PROGRAMS

Ireland

Bangladesh

Italy

South Africa

Vietnam

Mexico

Saudi Arabia

Turkey

Peru

India

Philippines

ThailandPakistan

CIVIL DNA DATABASESDiscussion for

whole population databases grows in

the Middle East

Denmark Study:“Nearly 80% say that cataloging the DNA of everyone in the country is a good idea.”- Copenhagen

Post(February 4, 2015)

Changing AttitudesKuwait

Oman

UAE

Decision Factors in

Choosing Loci:

THE LOCI USED FOR THE 60 MILLION SAMPLES AND FOR ALL SAMPLES IN THE IMMEDIATE FUTURE

Less focus on loci’s ability to deal with: Challenging Casework

Samples Mixtures Missing Persons/Mass

Disasters/War/Dead Familial Searching

<10 STR’sUK in 1995, other early adopters, parts of China

13-15 STR’sUSA and majority of other countries establishing databases after 2000

21-23 STR’sExisting gold standard

• What worked• Preventing

adventitious hits• Privacy• Time to result• Cost Consideration

If your goal is to get reliable hits against the database in these situations, what would you rather have in your reference database?

Degraded Casework, Mixtures, Missing Persons, Mass Disaster, War Dead, Familial Searching

Considerations/Drivers/Barriers:Enhanced Technology necessary to make it

practicalWhat markers should be used?Quantifying the positive impact on hitsPrivacy, policy, legal concerns

21-23 STRs or 21-23

STRs

Multiple Technologies Capillary Electrophoresis

Widely usedMultiplex analyses

Rapid DNA Typing InstrumentationCE basedProvides access by lay individuals

Microarrays Massively Parallel Sequencing

Multiplex analysesMultiplex samplesIncreased discrimination power

Multiple Technologies CE and MPS, in particular, offer the potential

to add more markers to the toolbox Current technology can meet needs Newer technologies provide long term

solutions Need discussions on markers that should be

considered to upload into a databaseWith consideration of country legislation

Current Forensic DNA Workflows

CE-based systems are the mainstay of DNA typingWell validated

○ Well understood○ Robust

Well establishedCost-effective on a per sample basisNo need to batch samplesSubstantial experienceExploit value of STRsResource investment already

in placeCan add more markers to

current format

Precedent for Expanded Marker Kits

Made Possible with 6-Dye Configuration

Expand on Theme

Increase number of dyes Allow for other markers to be

multiplexedSelection based on needs and identity

testing only Choice of additional markers

Markers for Possible Expansion

Those that allow better analysis of the bulk of caseworkNew STRs

○ Higher discrimination power○ Intra-allelic variants

Identity SNPsLineage markers

○ Haploblocks○ mtDNA○ Y STRs/SNPs

Types of SNPs Individual Identification SNPs:

SNPs that collectively give very low probabilities of two individuals having the same multisite genotype; individualization, High heterozygosity, low Fst

Ancestry Informative SNPs: SNPs that collectively give a high probability of an individual’s ancestry being

from one part of the world or being derived from two or more areas of the world Lineage Informative SNPs:

Sets of tightly linked SNPs that function as multiallelic markers that can serve to identify relatives with higher probabilities than simple di-allelic SNPs

Phenotype Informative SNPs: SNPs that provide high probability that the individual has particular phenotypes,

such as a particular skin color, hair color, eye color, etc. Pharmacogenetic SNPs – molecular autopsy

Indels Separated by size Fit well with CE format

Substantial Data on indels

The New Generation of Sequencing Technologies

First generation sequencing technology• Sanger Sequencing

Next generation sequencing technologies• Roche – 454• SOLiD• Illumina – GA II/HiSeq/MiSeq• Ion Torrent – PGM, Proton• Helicose• PacBio• Oxford Nanopore

Illumina MyGenome App

Illumina MiSeq™

Massively Parallel Sequencing

• Value• Backward compatibility with CE-based STR data• Large battery of genetic markers can be analyzed

simultaneously • Autosomal STRs, Y STRs, X STRs, and SNPs

(hundreds of markers)• mtDNA • Barcoding 16 to 384 (in theory) – multiple

individuals • Economies of scale

• Can be cost effective on a per marker basis

STR Panel

Locus AlleleNumber of

Varying Sequences

vWA 14 2

vWA 15 2

vWA 16 2

D3S1358 15 3

D3S1358 16 3

D8S1179 12 2

D8S1179 13 3

D8S1179 14 2

Mixture

Locus Repeats Coverage Sequence

D2S441 11 727 TCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATCTA

D2S441 14 664 TCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATTTATCTATCTA

D2S441 15 516 TCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATCTATTTATCTATCTA

D2S441 10 356 TCTATCTATCTATCTATCTATCTATCTATCTATCTGTCTA

D2S441 10 80 TCTATCTATCTATCTATCTATCTATCTATCTATCTATCTA

Allele 10 from one contributor and stutter (same length of a 10 allele with 80X coverage) from the 11 allele of the other contributor of the two-person mixture.

9 10 11 12 130

1000

2000

3000

4000

5000

6000

D5S818

Major Shared Minor

Nominal Alleles by Repeat

Dept

h of

Cov

erag

e

Minor and Major Contributor Alleles

Mixture of 2 peopleBoth 11,12

9 10 11 12 130

1000

2000

3000

4000

5000

6000

D5S818

Major Shared Minor

Nominal Alleles by Repeat

Dept

h of

Cov

erag

e

11 indistinguishable

Stutter from allele 11

12 distinguishable

AGAT...AGAT

AGAT...AGAT

AGAT...AGAG

Stutter from allele 12 of the minor

contributor

STR Selection Criterion New STRs with intra-allelic variation Identifying those markers with more alleles (ideally with similar

distributed allele frequencies) can seem sufficient for CE However, markers containing intra-allelic SNPs that display

similarly high heterozygosity are more desirable long term for forensic purposes

Length only approach translates into greater number of alleles with a large size difference Heterozygotes can result in substantial preferential amplification of the

smaller sized allele (or drop out of larger allele) A similarly discriminating locus due to the presence of intra-

allelic SNPs could have fewer examples of large size difference for heterozygote alleles Thus may demonstrate less preferential amplification

Privacy, Legal and Legislative Issues – New Technology

CE Technology Enhancements

MPS – Technology

Admissibility processes

Will MPS be permitted in a compulsory DNA environment?

○ Legislation may develop to highly regulate and restrict MPS from being in the hands of police based government agencies

○ Compare the Stingray Tracking Device https://www.aclu.org/map/stingray-tracking-devices-whos-got-them

Additional STRs Mito Identity SNPs Phenotypic and Ancestry SNPs Y-STRs

Privacy, Legal and Legislative Issues – Databasing additional STRs, Mito, ID SNPs,

Phenotypic SNPs, Ancestry SNPs and Y-STRs

• SNPs with high heterozygosity will not convey significant information about the variations for a Mendelian disorder even if there is complete linkage disequilibrium

• Very low predictive power

Identity SNPs, cont.

Privacy, Legal and Legislative – Phenotypic SNP’s and Ancestry Markers for Criminal Offender Databasing

Little utility to database phenotypic SNPs. Not practical on a cost benefit basis.

IrisPlex: Walsh et al. (2011)

Existing legal restrictions my already be in place. May not be permitted under USA statute for

criminal offeders○ DNA information must be related to a law

enforcement identification purpose - 42 U.S.C. 14132 (A) (b) (3) (A)

May not be permitted under USA Constitution ○ Skinner v. Ry. Labor Executives Assocsaiton (1989)

– “Physiological data is a further…invasion of privacy interest”

Privacy, Legal and Legislative – Phenotypic SNP’s and Ancestry Markers for Criminal Offender Databasing

Legislation likely to regulate the use of phenotypic and ancestry SNPs, ○ “Appearance” SNPs – Is your race, hair and eye color private? ○ Might be distinction between ancestry SNPs and SNPs that encode

facial morphology/pigmentation○ Some AIMs might be associated with disease genes

Requires more effort to determine if there are associations

○ “Sensitive” SNPs (disease, etc.)

Will USA Constitution allow for phenotypic and ancestry testing for casework? ○ Traits Exposed to the Public: (USA v. Mara – 1973)○ Abandonment concepts: California v. Greenwood, 486 U.S. 35 (1988)○ Abandonment applied to DNA: State v. Athan, Supreme Court of

Washington, 5/10/07 – Limited to confirming Identity ○ What will the court do with phenotypic and ancestry SNPs for casework?

Unlike STRs, will likely need a warrant.

Privacy, Legal and Legislative – Phenotypic, Ancestry SNP’s for CASEWORK

Privacy, Legal and Legislative – Multiple Y-STRs for Criminal

Offender Databasing Impact: Familial searching demand will increase

Consequence: Privacy fears might rise and legislation will increase to regulate familial searching

Recommendations to allow familial searching to continue:Understand and accept opposition’s concerns as a valid

point of viewSupport legislation to limit use, define protocols, and

provide for penalties for abuseDevelop model protocols for effective use with reduced

privacy intrusion

Identical Twins

Identical Twins

Little issue of the SNPs that may differentiate identical twinsUnlikely to provide privacy concerns (low predictive

power) No guidance on what to do with whole genome

dataDestroy data that are identicalProtective order on data disclosure

○ Criminal punishment

Defense right to review all data

Conclusions Casework is changing Different types of markers may accommodate better

the changing landscape of forensic evidence Technologies exist that enable an increase in the core

markers STRs, Identity SNPs/indels – unlikely to have privacy

concerns Phenotype and ancestry markers should not be

considered for entry into a DNA database Y STRs (and mtDNA) could assist in familial searching Need to consider legislation and/or model protocols for

effective use