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34 Cards in this Set
- Front
- Back
- 3rd side (hint)
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what is the name for the enzymes responsible for cutting DNA for recombinant techniques?
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restriction endonucleases
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what is unique about the recognition sequences typically recognized by restriction endonucleases?
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they are palindromic
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toward what end of the gel does DNA run?
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anode
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which cuts more frequently: a 4-, 6-, or 8-base pair endonuclease?
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4-base pair
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which migrate faster toward ends of gel, smaller or larger DNA fragments?
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smaller
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which enzyme is used in recombinant DNA technology to covalently seal DNA strands when they come together in the test tube?
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ligase
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how is vector DNA replication synchronized with the replication of the bacterial genome?
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it is not; vector replication is autonomously replicating DNA.
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vectors that carry foreign DNA into bacterial cells must be in which 2 forms?
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plasmids or bacteriophages
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what are 2 ways a vector can incorporate the foreign DNA into it?
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replace a fragment with the foreign DNA or add foreign DNA to their own sequence
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what are plasmids?
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double-stranded circular DNA elements
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where is the origin of replication for a plasmid?
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on the plasmid itself -- they have their own ORI
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what is the size limit of foreign DNA that a plasmid can accommodate?
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10kb
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what genes are necessary in a plasmid?
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antibiotic resistance genes
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what is a bacteriophage?
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type of DNA virus that infects bacteria
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None
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through which life cycles do bacteriophages replicate?
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lytic
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which portion of the phage genome can be replaced by the foreign gene?
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the segment devoted to the lysogenic life cycle
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what is the limit to the amount of foreign DNA a phage can accommodate?
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40kb
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in a field of bacteria, how is a phage recognized?
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as a plaque where all surrounding bacteria have been lysed
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what are the names for 2 techniques that can accommodate foreign DNA over 50kb? What is the major disadvantage for these techniques?
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cosmids and bacterial artificial chromosomes; they are very slow
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what are the two different DNA clones?
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genomic DNA clones and cDNA clones
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a genomic DNA clone represents what kind of DNA?
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segment of genomic DNA that may contain any element of the genome
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what is the normal size of a gene?
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over 30kb
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in which vectors are genomic DNA clones normally carried? Why?
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bacteriophages or cosmids or BAC's; because they are too big for plasmids
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for what does cDNA stand?
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complementary DNA
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why is cDNA created using mRNA?
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because mRNA does not include introns, promoter, or intragenic sequnces.
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what is a major difference between cDNA and genomic DNA?
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cDNA has no introns, promoters, or intragenic sequences.
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from which type of cells can genomic DNA be isolated? cDNA?
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any cells; cells specific to the expression of mRNA complementary to the cDNA
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are cDNAs normally shorter or longer than genomic DNA? Why?
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shorter; they have no introns
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which enzyme is necessary to create cDNA from mRNA?
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reverse transcriptase
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what is the sequence that can be used as a primer for reverse transcriptase in production of cDNA? Why?
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poly-T; complementary to poly-A tail of mRNA
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how is mRNA degraded in the production of cDNA?
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RNAse H
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how is a second strand of DNA created for cDNA?
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DNA polymerase
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what is one way that primers are created for production of 2nd DNA strand in cDNA?
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RNAse H leaves RNA primers behind via limited cleavage
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what is the final step in cDNA creation following prodcution of a double-stranded DNA molecule?
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addition of short linkers containing specific restriction sites at each end and the cDNA inserted into cloning vector and propogated in bacteria
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