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DNA Fingerprinting of Mycobacterium bovis by Restriction Fragment Length Polymorphism (RFLP)
What is RFLP?
How is DNA "fingerprinting" by RFLP performed?
Figure 2 provides a schematic diagram of the steps involved in producing a RFLP "fingerprint."
Step 5. When X-ray film is exposed to the nylon membrane, light produced from the probe will produce a banding pattern on the film. The number and location of bands will correspond to the original number and position of the insertion sequence in the bacteria.
How is RFLP used?
M. bovis reproduces by dividing itself in two. The genetic makeup of the new cells are identical to the original parent cell. Even after many generations, bacteria that have been produced from the original parent will share virtually all of the same genetic material (DNA) of the parent. The "fingerprints" of genetically related strains of M. bovis will be identical. Unrelated strains can be easily distinguished since their "fingerprints" will be different.
RFLP analysis is used by laboratory scientists and epidemiologists to evaluate the genetic relationship between isolates of M. bovis.
An example of PGRS patterns comparing M. bovis isolates is shown in figure 3.
The caption for figure 3: RFLP results when DNA is digested with AluI and probed with the probe PGRS. Lane 1, deer isolate; lane 2, raccoon isolate; lane 3, raccoon isolate; lane 4, coyote isolate; lane 5, coyote isolate; lane 6, coyote isolate; lane 7, coyote isolate; and lane 8, MI cow isolate.
The patterns of the deer and cow isolates, which resemble bar codes, are identical to each other. This indicates that the M. bovis isolates from the cow and the deer are related to each other.
Figure 4. RFLP results when DNA is digested with PvuII and probed with IS6110-445 base probe. Lane 1, deer isolate; lane 2, raccoon isolate; lane 3, raccoon isolate; lane 4, coyote isolate; lane 5, coyote isolate; lane 6, coyote isolate; lane 8, MI cow isolate.
The MDCH Molecular Biology Section performs RFLP analysis of both M. tuberculosis. and M. bovis.
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