The following simplified example demonstrates how restriction enzymes can be used to create a physical map of the location of restriction sites within genes on a strand of DNA.
This example describes an experiment where a single strand of DNA is analyzed to determine the position of 7 restriction sites, and the resulting gene fragments are arbitrarily labeled A, B, C, D, E, F, and G. It is assumed that a collection of restriction enzymes (abbreviated RE) are available that can cleave this strand at various locations within the gene, and that appropriate probes are available to analyze the fragments obtained from gel electrophoresis. (In practice, selective probes would be used so that not all fragments would be detected. In addition, it is highly unlikely that each fragment would be the same size.)
The following results are obtained from a series of experiments. The goal is to determine the correct gene fragment sequence. The table below shows the results obtained from a series of gel electrophoresis experiments. The DNA section of interest was cloned using either virus/bacteria growth or by the PCR. Samples were then cleaved with restriction enzymes to produce DNA fragments. These DNA fragments were separated using gel electrophoresis and analyzed with specific probes. The gene fragments present in each DNA fragment are shown in the table below.
| R.E. #1 | R.E. #2 | R.E. #3 | R.E. #4 |
|
. . . * BDEG * ACF . . |
* ABDEFG . . . . . * C |
. . . * BDFG . * AC * E |
. . . * ACFG . * BE * D |
|
Start of Gel Electrophoresis at top. Larger fragments move more slowly |
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Last modified March 12, 1997
Kent State University - Stark Campus
Department of Chemistry
This Page Written and Maintained by Dr. Clarke Earley