Using a blot test? A scientist at the San Diego Zoo is sent DNA samples from new

Question

Using a blot test? A scientist at the San Diego Zoo is sent DNA samples from newly isolated bacteria to analyze for a genomics study. The samples were collected from two different species isolated in different parts of the world. The labels have rubbed off of the tubes, however, so she does not know which sample is which. One of the organisms lived deep in the ocean at temperatures around 10 degree C and the other was isolated from a salt marsh in Death Valley, where temperatures can reach 45 degree C. How could she determine the most likely identities of the samples? Provide a specific reason why she can with some confidence determine the identities of each sample.

Explanation / Answer

The bacteria which is isolated from the deep ocean(temparature around 10 degree centigrade) are termed as psychrophiles whereas the bacteria which is isolated from the Death valley (temparature 45 degree centigrade) are called thermophiles.

In order to identify the samples the scientist can perform Southern blot hybridization technique.

First she should subject the sample DNA (bacterial DNA) to digestion by restriction endonucleases.This will produce short cleaved DNA fragments which can be seperated on the basis of size on agarose gel.Next, these seperated double stranded DNA fragments are denatured to yield single stranded DNA strands and these single stranded DNA strands are transferred to a nitrocellulose membrane.

Next, she need to prepare a labeled hybridization probe that is a DNA probe in nature.The label can be an antibody based marker.For the two bacterial species she can use two different labels that is two different antibody based markers. The probe will hybridize to target DNA sequence which is present in the sample DNA(bacterial DNA) due to complementary base pairing between them.

Next she will have to denature the probe in order to produce single stranded DNA probe.Denaturation of the probe can be done in various ways like application of alkaline conditions or by heat.This single stranded denatured DNA probe will base pair or hybridize with the single stranded target sequence in sample DNA.

Now, she will have to treat the nitrocellulose membrane with the probes and the probes which will find complementarity with the target DNA sequence will hybridize with it in the membrane.

Next,for visualization she need to to raise antigens against the antibody labels so that they can bind to the antibody based marker label on the probe.Since she used two different antibody based markers therefore after visualization she can distinguish the two species.

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