The prospect for more than 13 commonly used str tetranucleodide loci in forensic settings
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The prospect for more than 13 commonly used STR Tetranucleodide loci in forensic settings. Carrie Rowland Forensic Bioinformatic Services, Inc. Dayton, Ohio www.bioforensics.com. An Overview : Where are we going. History of DNA testing Generations of DNA tests and how performed

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The prospect for more than 13 commonly used STR Tetranucleodide loci in forensic settings

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The prospect for more than 13 commonly used str tetranucleodide loci in forensic settings

The prospect for more than 13 commonly used STR Tetranucleodide loci in forensic settings

Carrie Rowland

Forensic Bioinformatic Services, Inc.

Dayton, Ohio

www.bioforensics.com


An overview where are we going

An Overview: Where are we going

  • History of DNA testing

  • Generations of DNA tests and how performed

  • Current methodology

  • New prospects

  • Conclusions


Six criteria for judging a dna profiling system

Six criteria for judging a DNA profiling system

  • Discrimination power.

  • Sensitivity.

  • Absence of, or the ability to reconcile

  • technical artifacts.

  • Speed.

  • Ability to deconvolve mixtures.

  • Amenability to database searches.


Brief history of forensic dna typing

Brief History of Forensic DNA Typing

  • 1980 - Ray White describes first polymorphic RFLP marker

  • 1985 - Alec Jeffreys proposes multilocus VNTR probes be used forensically

  • 1985- first paper on PCR

  • 1988 - FBI starts DNA casework

  • 1991 - first STR paper

  • 1995 - FSS starts UK DNA database

  • 1998 - FBI launches CODIS database


Three generations of dna testing

Three generations of DNA testing

RFLP

AUTORAD

Allele = BAND

DQ-alpha

TEST STRIP

Allele = BLUE DOT

AutomatedSTR

ELECTROPHEROGRAM

Allele = PEAK


How do they fare

How do they fare?

STR

DQ-alpha

RFLP

 Discriminating power

Sensitivity

□ Technical artifacts

 Speed

□ Mixtures

 Databasing

 Discriminating power

□ Sensitivity

□ Technical artifacts

□ Speed

□ Mixtures

ٱDatabasing

□ Discriminating power

 Sensitivity

Technical artifacts

 Speed

□ Mixtures

□ Databasing


The prospect for more than 13 commonly used str tetranucleodide

RFLP technology

What we would

like for them to look like.


The prospect for more than 13 commonly used str tetranucleodide

What they often

look like.

Incomplete digest


Available kits for str analysis

Available Kits for STR Analysis

  • Kits make it easy for labs to just add DNA samples to a pre-made mix

  • 13 CODIS core loci

    • Profiler Plus and COfiler (PE Applied Biosystems)

    • PowerPlex 1.1 and 2.1 (Promega Corporation)

  • Increased power of discrimination

    • CTT (1994): 1 in 410

    • SGM Plus™ (1999): 1 in 3 trillion

    • PowerPlex ™ 16 (2000): 1 in 2 x 1017


An example forensic str multiplex kit

AmpFlSTR® Profiler Plus™

Kit available from PE Biosystems (Foster City, CA)

200 bp

300 bp

400 bp

100 bp

Size Separation

D3

vWA

FGA

5-FAM (blue)

JOE (green)

A

D8

D21

D18

Color Separation

D5

D13

D7

NED (yellow)

ROX (red)

GS500-internal lane standard

9 STRs amplified along with sex-typing marker amelogenin in a single PCR

An Example Forensic STR Multiplex Kit


The prospect for more than 13 commonly used str tetranucleodide

13 CODIS Core STR Loci with Chromosomal Positions

TPOX

D3S1358

TH01

D8S1179

D5S818

VWA

FGA

D7S820

CSF1PO

AMEL

D13S317

AMEL

D16S539

D18S51

D21S11


Str allele frequencies

45

40

35

30

Caucasians (N=427)

25

Frequency

Blacks (N=414)

20

Hispanics (N=414)

15

10

5

0

6

7

8

9

9.3

10

Number ofrepeats

STR Allele Frequencies

TH01 Marker

*Proc. Int. Sym. Hum. ID (Promega) 1997, p. 34


The prospect for more than 13 commonly used str tetranucleodide

Human Identity Testing with Multiplex STRs

AmpFlSTR® SGM Plus™ kit

DNA Size (base pairs)

D3

TH01

amelogenin

D8

VWA

D16

D19

D21

D18

D2

FGA

probability of a random match: ~1 in 3 trillion

Two different individuals

Results obtained in less than 5 hours with a spot of blood the size of a pinhead

D3

amelogenin

D19

D8

D16

VWA

D21

D18

D2

FGA

TH01

Simultaneous Analysis of 10 STRs and Gender


Fbi s codis dna database

FBI’s CODIS DNA Database

Combined DNA Index System

  • Launched October 1998

  • Used for linking serial crimes and unsolved cases with repeat offenders

  • Links all 50 states

  • Requires >4 RFLP markers and/or 13 core STR markers

  • Current backlog of > 600,000 samples

    As of June, 2004

  • Total profiles = 1,857,093

  • Total forensic profiles = 85,477

  • Total convicted offender = 1,771,616


The prospect for more than 13 commonly used str tetranucleodide

http://www.fbi.gov/hq/lab/codis/aidedmap.htm


Str alleles with n 4 stutter products

6.3%

6.2%

5.4%

STR alleles with n+4 stutter products

DNA Size (bp)

D8S1179

D21S11

D18S51

Relative Fluorescence Units


Schematic of stutter product formation process

1

2

3

4

5

6

4

5

1

2

3

Repeat unit bulges out when strand breathing occurs

Schematic of Stutter Product Formation Process

Normal STR Allele Replication

Slipped Strand Mispairing Model

Walsh et al (1996) Nucleic Acids Res. 24: 2807-2812


Mechanism behind n 4 stutter

Mechanism behind n+4 stutter

DNA polymerase

7

1

2

6

3

5

4

4

3

5

2

6

1

Template DNA

Walsh, et al. (1996) Nucleic Acids Research 24(14):2807


What is on the horizon

What is on the horizon ?

1. 14th …….locus

2. Pentanucleotide repeats

  • Mitochondrial DNA

  • Single nucleotide polymorphisms (SNPs)


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