BIO-RAD Forensic DNA Fingerprinting Kit

Section Concepts: DNA Scissors Activity, restriction enzymes, gel electrophoresis, standard curve

Background Information

Restriction enzymes: found in bacteria and archaea, are enzymes that cuts DNA at specific recognition nucleotide sequences known as restriction sites.

Gel Electrophoresis: a method used in biochemistry and molecular biology to separate a mixed population of DNA and RNA fragments by length, to estimate the size of DNA and RNA fragments, or to separate proteins by charge. Nucleic acid molecules are separated by applying an electric field to move the negatively charged molecules through an agarose matrix.

Restriction Enzymes + Gel Electrophoresis

Restriction enzymes video

Gel Electrophoresis animation

Timeline/Overview

Monday: Lecture Tuesday: Activities 1 & 2, DNA scissors,

pre-lab quiz Thursday: DNA fingerprint (restriction

digest), Analyze lab (work on packets) Monday: No school Tuesday: DNA fingerprinting kit (run gel,

visualize) Thursday: Packet due, post quiz

Necessary Preparation for Lab DNA Finger printing

DNA fingerprinting Labels

• 8—clear 1.5mL tubes ENZ (enzyme)• 8—clear 1.5mL tubes M (marker)• 8—clear 1.5mL tubes LD (loading dye)• 8--green 1.5mL tubes CS• 8—blue 1.5mL tubes S1• 8—orange1.5mL tubes S2• 8—violet 1.5mL tubes S3• 8—red 1.5mL tubes S4• 8—yellow 1.5mL tubes S5

Before Digest• Set water bath for 37C• Rehydrate & aliquot (page 13)

• EcoRI/PstI enzyme mix (12 hours before use)• Crime scene DNA • Suspects 1-5 DNA

Before Running Gel• Prepare electrophoresis TAE buffer• 11/13 at before 5th period Pour 8 gels (1%, EtBr, 10 well comb)• Prep. HindIII standard (right before class)• Aliquot

• HindIII (marker)• Loading dye

DNA Fingerprinting Lab - Background Information

DNA Fingerprinting: technique used by scientists to identify an individual based on their specific DNA profile

Restriction Digestion of DNA: restriction enzymes recognize particular sequences in a segment of DNA and cuts the DNA molecule at that point

Plasmids: circular, non-chromosomal pieces of DNA that can replicate in and are commonly found in bacteria

DNA Fingerprinting

DNA Fingerprinting Video

Plasmids

Restriction Enzyme Digestion of Plasmid DNA

Restriction Enzyme Animation

Restriction Enzyme EcoR1 Animation

DNA Fingerprinting Tips

Place tubes in refrigerator after removing from water bath

Gel will be 10 wells, adjust lanes in your protocol to use 2-8