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wrote...
13 years ago
Describe the use of a DNA probe and PCR for : (a) rapid identification of an unknown bacterium and (b) determining which of a group of bacteria are closely related.
The GC content of Micrococcus in 66-75 moles %, and of Staphylococcus , 30-40 moles %. According to this information, would you conclude that these two genera are closely related? Please explain your answer.
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Valued Member
13 years ago
You extract and PCR your bacterium's DNA, 4 cycles will do. You send in a restriction endonuclease (Restriction enzyme) that makes blunt cuts to make it linear (i.e. bacterial dna is cicular). Then you warm it up to 95°C and then lower it to 65°C to make it single stranded.

Now here's the kicker. If you have a probe library of known bacteria, grab a 96 well microsatellite plate and put your different known probes in each well, then add your your unknown bacterium DNA in each well and see where it binds fully, because the SSdna of the probe and the linear SS dna of the bacterium, if correspondant to each other, they will bind to make DSdna and your well will kight up in UV light (given your probes have some signalling method, be it radioactive or colorimetric).

This is done routinely in biotech labs and takes about 2 hours to identify an unknown.



No, G-C 66-75 moles % compared to 30-40% doesn't seem very close. But the more important thing is that G-C content doesnt tell you really how closely related 2 things are. Its the sequence of bases (ie. A,T,G,C) that matters. Percent conservation of sequence will tell you how closely related to things are. I think (but dont quote me on this) that humans and ape DNA is about 98% conserved. I'd say thats closely related.
wrote...
Valued Member
13 years ago
I have to say I've never seen moles written as a percentage. I would conclude from that information that Micrococcus and Staphylococcus are not particularly closely related, but it's hard to get much out of GC content alone. Take, for instance, E.coli, Shigella and Salmonella, three very closely related bacteria. They each have roughly 50% GC content. Take a 'few' steps away to Haemophilus, and it's at rougly 38%. Imagine the sheer number of mutations that would need to occur to significantly change the GC % of a cell with a few million base pairs. Try a google search on phylogeny "GC content" bacteria evolution, should give you something. At any rate, GC content is just one tool in bacterial phylogenies. A very useful one in particular with lateral gene transfer IDs.
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