AT Genetics and Molecular
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49 terms
Terms | Definitions |
|---|---|
 Recombinant DNA technology | Researchers use ______ to carry out five basic operations |
| gel electrophoresis | Cut enormously long strings of DNA into much smaller fragments with restriction enzymes and separate them by _____ |
| molecular cloning | Isolate, amplify, and purify fragments through_____ |
| hybridization probes | Use purified DNA fragments as___to identify similar sequences in libraries of clones or mixtures of DNA or RNA molecules |
| Polymerase Chain Reaction | Rapidly isolate and amplify previously defined genomic or mRNA sequences through the ____ |
| sequence of bases | Determine the precise___within isolated DNA fragments |
| restriction enzymes | ___fragment the genome at specific sites |
| blunt or sticky end | Restriction enzymes cut DNA molecules at specific locations and produce either ____ |
| blunt end | no hangover |
| 5' sticky end | 5' overhangs |
| 3' sticky end | 3' overhangs |
| size | Gel electrophoresis separates DNA fragments according to___ |
| Agarose prep | 1. attach comb in clear acrylic plate 2. pour heated molten agarose into plate and cool 3. remove gel from plate, wells are formed |
| load gel and electrophoresis | 4. micorpipette is used to load each well 5. add of power supply from negative to positive from about 1hour to 20 hours |
| Visualize DNA fragments | 1. remove gel from gel box 2. expose gel to uv light 3. red bands that appear can be compared to size markers |
| Polyacrylamide gels | _____separate from small fragments of DNA |
| agarose gels | ___separate larger fragments |
| genomes | __of animals, plants, and microorganisms are too large to analyze using simple techniques such as gel electrophoresis and restriction mapping |
| cloning | ___is a means to identify a specific DNA fragment within the genome, purify away all of the other fragments, and amplify it, making many identical copies of the fragment |
| restriction mapping and DNA sequencing | cloning fragment can be analyzed by ____ |
| PCR | Purification and amplification of previously sequenced genomic regions |
| Molecular cloning | Purification and amplification of previously uncharacterized DNA |
| Molecular cloning summary | Cut DNA and insert fragments of specific sizes into vectors.Transport vector-insert molecules into living cells that make many copies.DNA clones are any amplified set of purified DNA molecule |
| ligation | _____of fragments into cloned vectors creates recombinant DNA molecules during this cloning step |
| sticky ends | ____facilitate recombinant DNA fabrication during this cloning step |
| vector and insert DNA | Cutting the vector and DNA fragments generates complimentary sticky ends that increase the efficiency of ligation between the_____ during this cloning step |
| vector-insert recombinants | Host cells take up and amplify ____ |
| transformation | vectors carry insert DNA into cells during this step of cloning |
| suspension of competent E. coli. | during transformation recombinant DNA molecules are added to a_____ |
| electroporation | Cells are heat-shocked or shocked with a high-voltage electric shock called______ during transformation |
| ampicillian resistance | cellular clones are colonies that represent each viable plasmid containing bacterial cell contain______ which allows for tranformant identification |
| b-galactosidase selection | Cells that turn blue DO NOT have an insert. b-gal gene is intact. Cells that are white DO have an insert. b-gal gene is interrupted by insert |
| centrifugation | Plasmids and inserts are separated from bacteria and vectors by____, restriction digests and gel electrophoresis |
| exponential | PCR (polymerase chain reaction) achieved____accumulation of target DNA |
| 20bp | Based on previously determined DNA sequence, develop short oligonucleotides (___)complementary to sequences flanking the target DNA. |
| Oligonucleotides | ____act as primers to copy DNA similar to DNA replication |
| left primer | 5' to 3' |
| right primer | 3' to 5' |
| target region | between left and right primer |
| doubles | Each cycle of replication ___ amount of target DNA. |
| Taq DNA polymerase and 4 deoxynucleotide triphosphate | PCR 1: purify and denature DNA from target source Added to solution containing two oligonucletide primers, _____ and ___ |
| base pair | PCR 2: primers ___ at sites flanking target sequence of genomic DNA |
| polymerization | PCR 3: ___from primers along templates |
| 94 C, 5, 20 | 1. ___ for _minutes (first round) or for __ seconds (subsequent rounds) |
| 50-60 C, 2 | 2. ___for __ minutes |
| 72 C, 2-5 | 3. ___ for __ minutes |
| Exponential amplification | 1. denaturs strands 2. base pairing of primers 3. polymerization from primers along template |
| use for PCR | genetic mapping, genotype detection, analyze traces of partially degraded DNA, Evolutionary studies and diagnosis of infectious diseases |
| evolutuionary studies | compare homologous sequences from related organisms, compare sequences from a variety of sources, studies of gene diversity |
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