Southern Blot analysis of DNA
The left panel is an electrophoretic gel stained with ethidium bromide. Total DNA has been extracted from nine bacterial DNA clones (lanes ##1-9), digested with a particular restriction endonuclease, and separated by electrophoresis. Starting about one-third of the way from the top, a bright smear is present, indicating the presence of a random collection of restriction fragments of various sizes. There are no discrete bands, as no one DNA sequence is present in more than one copy. The lane marked (L) is a molecular weight standard (a "Ladder"), with a series of DNA fragments at 100bp intervals. A cloned DNA fragment has been loaded in lane #10: its mobility corresponding to the four rung of the ladder indicates a size of about 400bp.
The middle panel is a
Blot autoradiogram of the same gel. The DNA in
gel is transferred ("blotted") to a filter. The filter is
then hybridized with a radioactively-labelled DNA (a "probe")
made from the DNA loaded in
lane #10. The filter is then exposed to X-ray
film. Where the probe DNA finds a complementary sequence in
blot, it base-pairs ("sticks") to that DNA. The radioactivity
exposes the film and produces a dark band on the X-ray film.
The right panel
is a schematic representation of the autoradiogram: the information
content is the presence or absence of bands, and the size of the
fragments. The bands in the autoradiogram show
DNA sequence homologous to the probe DNA
in clones ## 3, 4, & 8, with the expected size, and absent
clones ##1, 2, 5, & 9. [The probe sticks to itself in #10, as
This analysis indicates that the gene of interest has been successfully
cloned in the first set of plasmids, which can now be analyzed further.
Figure modified from © 2000 by Griffiths et al.; text material © 2009 by Steven M. Carr