Biology Protocols

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Restriction Fragment Mapping

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Restriction Fragment mapping is a simple analytical tool for determining unknown (cloned into a known vecctor) or confirming "known" DNA samples (cDNAs especially).

The components required depend on the DNA to be analysed, but require the use of at least two restriction enzymes. DNA to be analysed is cut individually by each enzyme and then together in a double digest. Using this method the number of sites present in the DNA for each enzyme can be determined, and then their relative position in the DNA sequence can be determined using the double digest.

In this way, using many restriction enzymes and pairs of those enzymes, unknown DNA can be analysed and characterised as to restriction sites present or known DNA samples can be checked for sites characterisitic to that specific DNA.

For example:

Restriction Fragment Map example

In the example above a 6Kb plasmid is cut with BamHI, EcoR1 and BamHI/EcoRI (double digest).

The digest with BamHI yields a single band, indicating one restriction site, linearising the plasmid (6Kb fragment)

The digest with EcoRI yields two bands, indicating two restriction sites (5Kb and 1Kb)

The double digest yields three bands (3Kb, 2Kb and 1Kb)

From this we can deduce the following map:

EcoRI------------(3Kb)-----------BamHI------------(2Kb)-----------------EcoRI---------(1Kb)--------------------

 


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