Question

9. On Worksheet 16.IIIB is a restriction map of bacteriophage lambda. You digest some lambda DNA with the enzymes BamHI and HindIII separately and then load the fragments into an agarose gel and perform electrophoresis. Next, you perform a Southern analysis using the 4,878-bp EcoRI lambda fragment as a probe. a. Draw a picture of the electrophoresis gel, using the outline of the stained electrophoresis gel in Worksheet 16.IIIB (the two smallest HindIII fragments will run off the gel.) b. Indicate which fragments will hybridize to the 4,878-bp probe. (Hint: Even if there is only a partial overlap of the probe with a restriction fragment, the probe will still hybridize to that fragment.) c. Draw what would be seen after the Southern analysis on the membrane in the second outline.

Answer 1

Shown below are restriction maps for bacterio- phage lambda, showing the BamHI, HindIII, and EcoRI sites. Fragment sizes are in base pairs. The gel and hybridization analysis outlines are provided for your use. BamHl map 5,505 bp 16,841 bp 5,626 bp 6,527 bp 7,233 bp 6,770 bp Hindill map 2,322 2,927 23,130 9,416 6,557 4,361 11 EcoRl map Probe 125 564 7,421 21,226 4,878 5,643 5,804 3,530 Stained electrophoresis gel BamHI Hindill Results of hybridization analysis BamHI Hindill Size, bp 24,000 -- 7 Sample well Sample 23130 16841 16,000 8,000 17223 165-67 = 5626 BamHI overlap to EcoRI 4878bp 5.56 4,000 2027 Hind III overlap to EcoRI 4878bp 2,000 2027 2322 bp