Chapter Eleven – Forensic DNA Analysis

Chapter Eleven – Forensic DNA Analysis

FRSC 8104Criminalistics II

Professor Bensley

FRSC 8104 – DNA Objectives1. Understand the basic principles of DNA

structure and analysis.2. Recall the history and major steps in the

development of Forensic DNA Analyses3. Describe and detail the methods of DNA

Extraction and Quantitation from an evidentiary sample.

4. Describe and detail the steps involved in the Polymerase Chain Reaction.

FRSC 8104 – DNA Objectives

5. Understand the importance of Short Tandem Repeats in Forensic DNA Analysis.

6. Understand the function of the CODIS system7. Describe various techniques used in the

visualization of amplified DNA.8. Convey the important concepts of advanced

DNA techniques to include Y-STR and Mitochondrial DNA analyses.

FRSC 8104 – DNA OutlineI. Introduction and Overview

A. Sir Alec Jeffreys (1985)1. Colin Pitchfork

B. Results reported in DNA analysis

FRSC 8104 – DNA OutlineII. Biological and Genetic Basics – VIDEO!

A. Basic DNA Principles

Nucleotide Structure (dATP)

FRSC 8104 – DNA OutlineB. Chromosomes, Genes, DNA Markers

Locus (plural loci) is the specific location of a gene or DNA sequence on a chromosome. A variant of the DNA sequence at a given locus is called an allele. The ordered list of loci known for a particular genome is called a genetic map.

FRSC 8104 – DNA OutlineC. DNA Marker NomenclatureD. Population Variation

SEQUENCE POLYMORPHISM LENGTH POLYMORPHISM

-------(AATG) (AATG) (AATG)--------3 repeats

------------(AATG) (AATG)------------2 repeats

III. Historical MethodsA. Pre 1985B. RFLP-Based DNA TypingRFLP AnimationSouthern Blot Animation

FRSC 8104 – DNA Outline

C. Early Based PCR Testing1. HLA DQ Alpha / DQA1

FRSC 8104 – DNA Outline

2. Polymarker (PM+DQA1)

FRSC 8104 – DNA Outline

3. D1S80

FRSC 8104 – DNA Outline

4. Short Tandem Repeats

FRSC 8104 – DNA Outline

D. Silver vs. Fluorescent Staining Techniques

FRSC 8104 – DNA Outline

FRSC 8104 – DNA OutlineIV. DNA Extraction

A. Organic ExtractionB. Chelex ExtractionC. FTA Paper

D. Solid Phase ExtractionE. Differential Extraction

FRSC 8104 – DNA OutlineV. DNA Quantitation

A. Why?B. How?

TAQMan

FRSC 8104 – DNA OutlineVI. DNA Amplification (PCR)

A. IntroductionB. Why PCR?C. Issues with PCRD. PCR Components

How PCR was Discovered

What the?!?!?

FRSC 8104 – DNA OutlineE. The Process of Replication

1. DNA Polymerase2. Okazaki Fragments

DNA Replication Process Video

http://rst.gsfc.nasa.gov/Sect20/DNA-20replication.gif

FRSC 8104 – DNA OutlineF. DenaturationG. AnnealingH. ExtensionI. Multiplex PCRJ. PCR Inhibition

Video of the PCR ProcessAnother Video of the PCR Process

FRSC 8104 – DNA OutlineVII.Short Tandem Repeat Markers (STRs)

A. Repetitious DNAB. Microsatellites or STR

FRSC 8104 – DNA OutlineC. 13 CODIS STR Loci

The Thirteen CODIS STRs and Their Probability of IdentitiesSTR African-American CaucasianD3S1358 0.094 0.075vWA 0.063 0.062FGA 0.033 0.036THO1 0.109 0.081TPOX 0.090 0.195CSF1PO 0.081 0.112D5S818 0.112 0.158D13S317 0.136 0.085D7S820 0.080 0.065D8S1179 0.082 0.067D21S11 0.034 0.039D18S51 0.029 0.028D16S539 0.070 0.089

Source: The Future of Forensic DNA Testing: Predictions of the Research and Development Working Group. Washington, DC: National Institute of Justice. Department of Justice, 2000, p. 41.

FRSC 8104 – DNA OutlineVIII.DNA Separation and Detection

A. Electrophoresis1. Slab Gels2. Capillary

B. DNA Detection

FRSC 8104 – DNA OutlineIX. Advanced Techniques

A. Y-STR

FRSC 8104 – DNA OutlineB. Mitochondrial DNA