Term
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Definition
| It amplifies DNA--makes you like billions of copies of a sequence in question. |
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Term
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Definition
| Polymerase chain reaction |
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Term
| What do you need for PCR? |
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Definition
| need a primer, DNA polymerase, nucleotides, PCR machine |
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Term
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Definition
| So, you need 2 primers complementary to the sequences at 3' end of gene of interest (one for template and one for non-template). They are about 16-20 nucleotides long, and then DNA synthesis occurs from there to the end of the gene. Then, the next round happens, and because you now have a complementary strand the other primer works and you get the actual size of the sequence you are looking for. |
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Term
| What are the steps of PCR? |
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Definition
| 90'c denaturing step. 50' annealing step (primers bind). 72' DNA polymerase extension step. Then run the cycles a bunch of times so you have tons of DNA of interest. |
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Term
| How could you use PCR to detect carriers of genetic defects? |
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Definition
| Run PCR to amplify gene of interest. Compare genes of wild-type and specimen with western blot. If size of gene is different, it is defective. Heterozygotes will have one normal and one defective. Homozygotes will only have defective. Wild-type will only have normal. |
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Term
| How could you detect foreign nucleic acid by PCR? |
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Definition
| Well, this would be useful for identifying infections. So, you'd have a primer for the sequence of interest in an organism of interest. Then, you'd run PCR and get lots of that DNA. The Primer would be special--fluorescence and quencher modification. |
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Term
| how does the fluorescence and quencher thing work with PCR? |
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Definition
| DNA polymerase will digest the fluorescent primer as it copies the bacterial DNA. This will release the quencher group and the fluorescence will be visible. Can detect as few as 5 organisms! Takes only about 7-9 minutes! |
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Term
| What is RT-PCR? Why is it special? |
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Definition
| You use reverse transcriptase to detect RNA by making DNA copies of it. It is special because you can quantify viral load by seeing how much DNA is made. It is very sensitive. |
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