1. DNA Fragment Analysis

2. DNA Fragment Analysis
What shows have you seen on TV that use or talk about DNA evidence?

4. How is DNA really analyzed?
Samples collected
DNA isolated and extracted
DNA is cut or amplified and sorted
-Most common method is gel electrophoresis
Results are analyzed

5. Collecting samples
Must be collected carefully to prevent contamination
Only a small amount needed

6. Isolating DNA
You isolated DNA from various plant and animal tissues the other week
Care must be taken to ensure DNA strand is not incomplete or damaged

7. Cutting or amplification of DNA
If the sample is large you can use restriction enzymes to cut the DNA into small pieces

8. Restriction Enzymes
Special enzymes that recognize specific sequences in DNA and cut it there
Name comes from the bacteria isolated from
Bacteria use them for defense

9. Restriction Enzymes Enzyme Organism from which derived
Target sequence (cut at *)5' -->3'
Ava I
Anabaena variabilis
C* C/T C G A/G G
Bam HI
Bacillus amyloliquefaciens
G* G A T C C
Bgl II
Bacillus globigii
A* G A T C T
Eco RI
Escherichia coli RY 13
G* A A T T C
Hae III
Haemophilus aegyptius
G G * C C
Hind III
Haemophilus inflenzae Rd
A* A G C T T
Hpa I
Haemophilus parainflenzae
G T T * A A C
Kpn I
Klebsiella pneumoniae
G G T A C * C
Mbo I
Moraxella bovis
*G A T C
Pst I
Providencia stuartii
C T G C A * G
Sma I
Serratia marcescens
C C C * G G G
SstI
Streptomyces stanford
G A G C T * C
Sal I
Streptomyces albus
G G * T C G A C
Taq I
Thermophilus aquaticus
T * C G A
Xma I
Xanthamonas malvacearum
C * C C G G G

10. Restriction Enzymes ….TGG ….ACC CCAGATGGCG GGTCTACCGCTTAA AATTCGGCG…
Eco RI
Hae III
….TGG
….ACC
CCAGATGGCG
GGTCTACCGCTTAA
AATTCGGCG…
GCCGC…

11. Cutting or amplification of DNA
If the sample is large you can use restriction enzymes to cut the DNA into small pieces
If sample is small copies of portions can be made
Polymerase Chain Reaction or PCR

12. https://fp.auburn.edu/asim/State BIOLOGY/Bio Labs/B8EleDye/Teacher/PCR_reaction.swf

13. Sorting fragments Process called gel electrophoresis
Gel made from agarose which is semi-solid
DNA fragments sorted by
Charge (DNA is negatively charged)
Size (base pairs)
Shape

14. Sorting fragments DNA fragments sorted by
Charge (DNA is negatively charged)
Size (base pairs)
Shape
How do you think these factors affect movement?

15. Electrophoresis (-) (+) DNA fragments Negative electrode
Positive electrode
(+)

16. Sorting fragments Small fragments travel further than large fragments
Bands are formed by large numbers of similar sized fragments
Standard or ladder is added to compare fragment sizes

17. Analysis Similar bands indicate similar DNA
Remember where your DNA comes from
Mom and Dad
Multiple bands are needed for positive identification

18. Review DNA fragments can be used to identify individuals
DNA is isolated, cut or amplified, and finally sorted by electrophoresis
DNA bands can then be analyzed