detección de moléculas específicas de adn, arn y proteínas

8/10/2019 Deteccin De Molculas Especficas de ADN, ARN y Protenas

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DETECCIN DE MOLCULAS ESPECFICAS DE ADN, ARN Y PROTENAS.

Because the main macromolecules of genetics are DNA, RNA and protein, genetic

analysis often requires the detection of specific molecules of each of these three types.

How can specific molecules be identified among the thousands of types in the cell? The

most extensively used method for detecting specific macromolecules in a mixture is

probing. This method makes use of the specificity of intermolecular binding, which we

have already encountered several times. The probe is labeled in some way, either by a

radioactive atom or by a fluorescent compound, so that the site of binding can easily be

detected. Let's look at probes for DNA, RNA, and protein.

Probing for a specific DNA

A cloned gene can act as a probe for finding segments of DNA that have the same or a

very similar sequence. For example, if a gene G from a fungus has been cloned, it might

be of interest to determine whether plants have the same gene. The use of a cloned

gene as a probe takes us back to the principle of base complementarity. The probe

works through the principle that, in solution, the random motion of probe molecules

enables them to find and bind to complementary sequences. The experiment must be

done with separated DNA strands, because then the bonding sites of the bases are

unoccupied. DNA from the plant is extracted and cut with one of the many available

types of restriction enzymes, which cut DNA at specific target sequences of four or

more bases. The target sequences are at the same positions in all the plant cells used,

so the enzyme cuts the genome into defined populations of segments of specific sizes.

The fragments can be fractionated by using electrophoresis.

Electrophoresis fractionates a population of nucleic acid fragments on the basis of size.

The cut mixture is placed in a small well in a gelatinous slab (a gel), and the gel is

placed in a powerful electrical field. The electricity causes the molecules to move

through the gel at speeds inversely proportional to their size. After fractionation, the

separated fragments are blotted onto a piece of porous membrane, where they maintain

the same relative positions. This procedure is called a Southern blot . After having

been heated to separate the DNA strands and hold the DNA in position, the membrane
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detección de moléculas específicas de adn, arn y proteínas

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