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Experiment 3:  Restriction Fragment
Length Polymorphism (RFLP)


As you may have learned, not all mutations are harmful. For instance, a single base pair change in a human gene is likely to effect the viability of the organism, although it is still technically a mutation. Single base changes like this occur very frequently and are part of life.

Now switch gears for a minute and think about Restriction Enzymes. If you're not comfortable with your knowledge about these proteins, see either Experiment 1 or the Background Information. Restriction Enzymes (RE's) are exquisitely specific in their action -- they will only cut the nucleic acid if the exact sequence is recognized. If even one base differs, the strand will not be cut. Take for example the figure below. In this figure, the sequence recognized by the RE EcoRI is shown (you may recognize the top part of the figure from the background information). If the exact sequence of GAATTC is found, the strands are cut by EcoRI at the sites indicated by the arrows. Now look at the bottom line of the figure. The only difference is the change of one base pair, shown in red. This strand will not be cut by EcoRI.

RFLP fig. 1

Is the change from a TA base pair to an AT base pair a mutation? Yes.

Will this mutation affect the organism that carries it? We can't say for sure without more information, but there's a good chance it won't.

Can we use this information to our advantage? Absolutely. This is the essence of RFLP analysis: my DNA will have accumulated different mutations like this than yours. Add to this the fact that at any given polymorphism site (a place where this type of difference occurs), one of your chromosomes may be different than the other, and you get a powerful tool not only in genetic counseling, but criminology as well.


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