Using RFLPs for mapping genetic diseases and for DNA fingerprinting

Restriction Length Fragment Polymorphisms

gene as a marker

1. sickle cell anemia


2. phenylketonuria

obtain a sample of blood, hair root, or other biological sample.

These fragments are generated by cutting genomic DNA with a restriction endonuclease at a particular nucleotide sequence and separating the resulting fragments on a gel by performing a Southern blot

(G^CGC)

These fragments are then run on an agarose gel in separate lanes and the fragments will migrate towards the positive electrode to different degrees based on the molecular weight of each fragment.

The smaller fragments will move farther on the gel than the larger fragments.

The fragments then need to be visualized. This is commonly done by transferring the bands to a nitrocellulose gel and probing for the various DNA sequences contained in the fragments.

Ideally, this probe is 6-10 bp, but in the simplified example above, the probe could be GCG, which would bind to the red CGC sequence contained by all fragments. The probe needs to be able to be visualized, and this can be done by exposing the radioactive probe to x-ray film.

At this point, the fragments of various lengths can be visualized and the sequence differences between these two people can be visualized. Person one will show 3 DNA fragments and Person 2 will show 2 DNA fragments.