PCR and Gene cloning

[FFK]

Hello I'm just starting to learn molecular biology. I hope someone out there able to explain the difference between gene cloning with plasmid and Polymerase chain reaction (PCR). Are they have the similar function (to amplify the DNA fragments) but work at different way? Thank you.


Regards,
Foo

[aftabac]

Hi
yes, both ways are used to amplify desired gene or DNA Fragement. but there is difference in application as by PCR DNA is amplified to analyze it but by plasmid cloning, DNA is stored for long time, analyzed and also used for expression in experiments. these are main difference, although there are planty of other differences.

regards
aftab

[FFK]

Thank you for sharing =). Neway, I've one more question here. Can we say microsatellite is the primer uses in PCR complementary with the known sequences? Thank you


Regards,
Foo

[hanami91]

PCR targets a segment of DNA with specially designed primers to ampifly it through a series of cyles in the thermocycler. The primers are typically @ 20 bases long and target the 5' end of each strand. In the first stage of a PCR cycle the strands are denatured, -heated (@94C) to disrupt the hydrogen bonds and they separate. In the second stage-annealing the temp lowers (@ 60-70C) to just below the primer melting point and the primers fix to their position on the DNA template. During elongation (@72C) the strands are elongated by the base supplied with the PCR reaction.

In gene cloning with plasmids, bacteria are transformed. Usually E. coli, to take up a desired plasmid which carries a desired genetic trait, and another trait used for selection such as a specific antibiotic resistance. The transformed bacteria are cultivated for a few days and then spread on petri dishes containing the antibiotic the plasmids are resistant to. Bacteria that grow on this have the plasmid. Colonies are then selected for pure cultures. From the pure cultures the plasmids can be extracted from the bacteria through miniprep.
Hope this was helpful,
haanmi91

[admin]

Thanks Hanami