Marker

1
1 Marker
2 Uncut
3 H
4 E
5 P
6 EH
7 HP
8 EP
Agarose Gel
Marker Size
Origin (wells)
21,000 bp
10,000
9,600 bp
Restriction Enzymes E EcoR I H Hind III P
Pst I
5,800
5,000
5,200 bp
4,800
4,300 bp
4,200
4,200
3,800
3,600
3,600
3,000
2,400
1,600
1,600
1,800
1,500 bp
1,600
1,200
1,300 bp
1,000
800
600 bp
1 Marker
6 EH
2 Uncut
3 H
4 E
5 P
7 HP
8 EP
Southern Blot 1 X-ray of southern blot probed
with 3 end gene
Origin (wells)
10,000
5,800
4,800
4,200
4,200
3,800
3,000
1,800
800
Southern Blot 2 X-ray of southern blot probed
with 5 end gene
1 Marker
6 EH
2 Uncut
3 H
4 E
5 P
7 HP
8 EP
Origin (wells)
10,000
5,800
5,000
4,800
1,800
1,600
1,600
1,600
1,000
2
Restriction Map Assignment

• Generate a restriction map based on the fragments
  observed in the Agarose Gel
• show all the enzyme cut sites and the number of
  base pairs between them, the shape of the DNA
  sample used, and the total size of the DNA used
• The plasmid contains a 2000 bp gene. Using the
  band sizes from the two southern blots, determine
  its location relative to the restriction map
  generated in step 1 (show the location of the
  gene on the restriction map). Assume that each
  probe corresponds to the first 200 bp of their
  respective ends (Hint Not all 200bp of your
  probe needs to bind to light up a band. Approx.
  50bp overlap is enough).
• Clearly indicate which end of the gene is 5 and
  which is the 3 end.
• Hand in the assignment, including your rough
  work, at the beginning of next weeks lab