1. PCR Y.Martinez, LSHS, 2014 DIRECTIONS: COPY NOTES IN ORANGE.

2. What is DNA profiling? DNA profiling is the use of molecular genetic methods to determine the exact genotype of a DNA sample in a way that can basically distinguish one human being from another. The unique genotype of each sample is called a DNA profile.

3. How do crime scene investigators create a DNA profile? 1. Evidence is collected at the crime scene: Blood Tissue Semen Urine Hair Teeth Saliva Bone

4. 2. DNA is extracted from sources at the crime scene and from victim and suspects

5. 3. DNA samples are processed Sample Obtained from Crime Scene or Paternity Investigation Biology DNA Extraction DNA Extraction DNA Quantitation DNA Quantitation PCR Amplification of multiple (STR) markers PCR Amplification of multiple (STR) markers Technology Separation and Detection of PCR Products (STR Alleles) Separation and Detection of PCR Products (STR Alleles) Sample Genotype Determination Genetics Comparison of Sample Genotype to Other Sample Results If match occurs, comparison of DNA profile to population databases Generation of Case Report with Probability of Random Match

6. 4. Crime Scene Investigators search in areas of the genome that are unique from individual to individual and are “anonymous” (control no known trait or function) The areas examined are Short Tandem Repeats or STR’s Humans are 99.9% so… STR region

7. To determine the genotype (DNA profile) Crime Scene Investigators make billions of copies of the target sequence using PCR. Starting DNA Template 5’ 3’ Target DNA

8. What’s the point of PCR?  PCR, or the polymerase chain reaction, makes copies of a specific piece of DNA  PCR allows you to look at one specific piece of DNA by making copies of *only* that piece of DNA  PCR is like looking for a needle in a haystack, and then making a haystack out of the needle

9. Exons and Introns *When DNA is made, exons and introns are made. *Exons are parts of DNA our body uses for everything! *Introns are non-coding regions of DNA or “Junk DNA”

10. PCR

11.  Suspects are included in an investigation if their DNA profile matches with genotypes found at the crime scene.  Suspects can be excluded if their DNA profile does not match genotypes found at the crime scene.  DNA profiling is never used alone; it is used in combination with other evidence.

12. What happens during PCR?

13. The PCR Reaction How does it work? The process occurs in a machine called a Thermalcycler. The Thermalcycler will go through a series of heating and cooling to activate enzymes and denature DNA. In about 3 hours, over 1 BILLION of copies of the desired gene is replicated.

14. PCR