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10 Cards in this Set
- Front
- Back
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What is a restriction endonuclease?
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A prokaryotic enzyme that cuts up foreign DNA (as in from a virus). Methylation of host bases provides protection from the enzyme.
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What are the 3 general class of vector?
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Plasmids
Phage Yeast artificial chromosomes (YACs) |
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What are plasmids?
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Small circular DNA molecules found in bacteria. Generally used to carry foreign DNA sequences
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What is a Phage?
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bacterial viruses.
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What is a YAC?
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Used to propagate foreign DNA molecules hundreds of Kbps in yeast cells. They utilize isolated centromere, telomere and origins of replication in yeast.
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What 3 features must a plasmid vector have to be useful?
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Origin of DNA replication
Selectable marker Unique restriction enzyme sites |
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How does cloning using a plasmid vector work?
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Foreign DNA is digested and along with the plasmid DNA with specific restriction enzymes.
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How does southern blotting work?
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DNA is cut into fragents using a restriction endonuclease. These fragments are run through electrophoresis to separate by molecular weight and a nitrocellulose sheet is laid over the gel. A radioactive probe is added containing a single nucleotide and autoradiography is done. Hybridization of the probe indicates that this fragment contains DNA complementary to the probe.
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How does PCR work?
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DNA is denatured at a high temperature. Primers are added and the temperature is lowered allowing the primers to anneal to the single strands of DNA. Taq polymerase and dTNP's are added allowing for polymerization of a new strand.
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What are the use of ddNTP's?
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They are dideoxynucleotides. Therefore, they lack both a 2' and 3' hydroxyl group and will terminate the growth of a chain. They are used to sequence DNA and is used in conjunction with electrophoresis.
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