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30 Cards in this Set
- Front
- Back
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What is DCPIP?
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An indicator that is originally blue in colour, but becomes clear when it is reduced.
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What is the lambda max for oxidized (blue) DCPIP?
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620 nm
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How can DCPIP be used to measure photosynthetic activity?
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DCPIP intercepts electrons travelling from photosystem II to photosystem I, it is reduced by gaining electrons, and looses its colour. The more molecules are reduced, the more lose their color, the less concentrated the colour will be.
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(photosynthesis experiment)
What are controls that need to be used? Why do we need them? |
-DCPIP only exposed to light (Some reduction may occur)
-DCPIP + chloroplasts kept in the dark (Some reduction may occur) -Chloroplasts only exposed to light (Some colour change may occur) |
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What are plasmids?
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Small, circular, self-replicating DNA molecules found in bacterial cells.
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What is the approximate length of a plasmid?
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100,000 to 200,000 base pairs
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What are plasmids currently used for?
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They are used as tools in biotechnology, mainly in the creation of genetically modified organisms.
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What is a vector in molecular biology?
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A DNA molecule used to carry foreign DNA sequences into the bacterium.
Plasmids are the simplest bacterial vectors. |
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Explain stringent control plasmids vs relaxed plasmids.
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Under stringent control: Only replicate with the main bacterial chromosome. Usually only one copy.
Relaxed: Replicate autonomously and have copy numbers of 10-500 per cell. |
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What plasmids are usually maintained in a bacterial population?
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Those that provide an importan selective advantage. Ex: Gene for antibiotic resistance.
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In nature, plasmids are passed on from one strain to another by conjugation, in a lab, this is done by transformation.
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In nature, plasmids are passed on from one strain to another by conjugation, in a lab, this is done by transformation.
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Why are plasmids so widely used in biotechnology?
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Their small size and ring structure makes them relatively easy to separate from chromosomal DNA
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What is a miniprep?
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A small preparation of purified plasmid DNA
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What is a recombinant plasmid?
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Modified plasmids containing foreign genes.
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What does SDS-sodium hydroxyde do in the context of Plasmid DNA purification?
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Acetic acid mixture kyses (degrades) the bacterial cells.
SDS disolves the lipid component of the cell envelope, as well as cellular proteins. Sodium Hydrxide denatures chromosomal and plasmid DNA into single strands. Plasmid DNA stay interwined. |
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What does Glucose-Tris-EDTA (GTE) do in the context of Plasmid DNA purification?
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Glucose maintains osmotic pressure.
Tris buffers the cell at pH 7.9 EDTA weakens the cell envelope (by binding divalent cations, has an affinity for divalent cations) After cell beraks down, EDTA prevents DNA degradation. (prevents formation of nucleases by binding Mg2+ ions, necessary cofactors in bacterial nucleases) |
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What does Potassium acetate-acetic acid do in the context of Plasmid DNA purification?
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Acetic acid returns pH to neutral (Allows DNA to renature)
Large DNA collapses in a tangle into the precipitate Potassium acetate precipitates the SDS and the proteins and lipids the SDS was associated with. As a result, only smaller DNA and RNA molecules remain in the solution. (They don't precipitate) |
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What does Isopropanol do in the context of Plasmid DNA purification?
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Rapidly precipitates nucleic acids, but slowly precipitates proteins.
A quick precipitation brings down nucleic acids. |
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What does Ethanol do in the context of Plasmid DNA purification?
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Wash with ethanol rmoves remaining salts and SDS from the preparation.
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What does Tris-EDTA (TE) do in the context of Plasmid DNA purification?
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Tris Buffers the DNA Solutons
EDTA protects the DNA from degradation by DNAses |
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What is another word for restriction endonucleases?
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Restriction enzymes.
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What are nucleases?
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Enzymes that can break down the phosphodiester bonds that link adjacent nucleotides in DNA and RNA.
Endonucleases: cleave at internal positions Exonucleases: progressively digest from the ends. |
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What type of restriction enzyme is most used in biology and why?
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Type 2 because:
- Only have restriction activity, methylation s done by different enzyme - Cut in a predictable and consistent manner at a site within or adjacent to the recognition sequence. - Do not require ATP as a cofactor, only Mg2+ ion. |
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Define Constitutive enzymes
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Produced at a constant rate under any conditions (almost). Ex: Enzyes of the central metabolic pathways, like glycolysis.
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Define Inducible enzymes
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Are present at very low levels, but concentration will rise if exposed to substrate. Substrate is called inducer in this case. Ex: Enzymes that broaden a cells ability to survive on different food molecules.
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Define repressible enzymes
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Usually present in significant concentrations on minimal medium, but disappear rapidly if exposed to a repressor. Ex: cells involved in anabolic pathways)
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What is the function of the LacZ operon?
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Codes for B-galactosidase, which hydrolyses lactose into glucose and galactose.
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What is the function of the LacY operon?
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Codes for galactoside permease, membrane protein which actively transports lactose and other galactosides into the cell.
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How does the operon work?
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In the absence of lactose, the operon is switched "off" by repressor bound to DNA. When lactose is present, it attaches itself onto the repressor, making it unable to stay attached to the DNA. Gene is turned n, producing shiz.
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How is IPTG important in the lac operon lab?
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IPTG is similar in structure to lactose, and thus it can turn on the operon.
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