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27 Cards in this Set
- Front
- Back
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Stress |
overpowering pressure of some adverse force or influence, biotic or abiotic, that tends to inhibit the normal functioning of a biological system pH, Temp., pressure, osmotic, UV, antibiotics, [O2 or CO2], [nutrients] |
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Responses to Stress |
death, sporulation, change in metabolism, dormancy, motility, apoptosis, membrane modifications |
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What does OD stand for? |
Optical density OD = 1 = 1 x 10^9 cell/ ml |
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What 2 organisms where used in the stress experiment? What were the 2 stresses that were tested? |
A prokaryote: E. coli A eukaryote: Saccharomyces cerevisiae (a yeast) Heat at 50 degrees Celsius and salinity |
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To what degree of a serial dilution did you create for the stresses experiment? |
Carried out a serial dilution of a culture to set up to a specific OD of 600 nm |
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What were the salt concentrations of the agar plates for salinity stress? |
Using dilution 10^-6 LB (E. coli): 0, 100, 250, 500 YPD (yeast): 0, 250, 500, 1000 E.c OD = 11.64 OD Y OD = 11.9 OD |
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What were the dilutions of yeast cells used for the heat stress? |
Serial dilutions of 10^-4 and -5 **in the presence of heat the number of colonies decreased on the plates; as the concentration of salinity increased the number of colonies also decreased** |
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What is the equation for computing the estimated number of bacteria on the surface of the plates? |
B = N/D B = number of bacteria N = number of colonies counted on a plate D = dilution factor 10^4 |
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Transformation |
the genetic alteration of a cell resulting from the uptake and expression of foreign genetic material (DNA) |
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Transduction |
genetic alterations resulting from introduction of DNA by plasmid, plasmid-encoded conjugation, or by viruses |
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Competent Bacteria |
Bacteria which are able to uptake DNA; 2 different types of competence: natural and artificial |
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Natural Competence |
some bacteria are naturally capable of taking up DNA under laboratory conditions; such species carry sets of genes specifying machinery for bringing DNA across the cell's membrane(s) |
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Artificial Competence |
Not encoded in the bacterial cell's genes; it is instead induced by laboratory procedures in which cells are passively made permeable to DNA, using conditions that do not normally occur in nature |
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Plasmid Transformation |
after the heat shock step intact plasmid DNA molecules replicate in bacterial host cells |
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What is the F-pilus in the infection of a bacteriophage? |
The F-pilus is the rate-limiting step in the infection The E. coli in the transduction experiment were starved to their [F-pilus containing bacteria] |
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What is the equation for determining the transducing units of a sample? |
TU = # colonies/ Dilution factor (# of virions/ ml) |
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Principle Information for Transformation/ Transduction |
Pelleted bacterial cells are resuspended and subjected to SDS/ alkaline lysis to: 1. liberate the plasmid DNA 2. the resulted lysate is neutralized to create appropriate conditions for binding of plasmid DNA on the silica membrane in the spin column The absorbed DNA is washed to remove contaminants, and is then eluted with a small volume of the Elution Buffer (10 mM Tris-HCL, pH 8.5) |
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Restriction enzymes |
DNA-cutting enzymes found in bacteria; recognize and cuts DNA only at a particular sequence of nucleotides; cut in an offset fashion **Require magnesium for activity, but they have different buffers for pH and salt concentrations |
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Process of a restriction enzyme (restriction endonuclease) cutting DNA |
Scans length of DNA molecule, recognizes a specific sequence, bonds to the DNA molecule, and makes one cut in each of the two sugar-phosphate backbones of the double helix |
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"Sticky ends"
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The ends of a cut, usually a overhanging piece of single-stranded DNA; able to form base pairs with any DNA molecule that contains the complementary end
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Types of enzyme cuts: |
5' overhangs: cuts asymmetrically, results in a single-stranded overhang extends from the 5' end 3' overhangs: asymmetrical cutting, results in a single-stranded overhang from the two 3' ends Blunts: cut at precisely opposite sites in the two stands of DNA generate blunt ends without overhangs |
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Isoschizomers |
different restriction enzymes can have the same recognition site |
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Electrophoresis |
permits one to see something directly related to the physical and informational properties of DNA Neg. DNA --> Pos. Electrode |
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Relationship b/t the size of DNA fragment and the velocity it travels through a gel complex? |
The log of the size of the fragment is more or less inversely proportional to the velocity of the fragment and hence to the distance traveled at the end point Lg. DNA = slow Sm. DNA = fast |
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Ethidium Bromide |
a dye that fluoresces orange when UV light shines on it, so DNA can be seen; used to monitor the progress of the electrophoresis |
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The GeneJET Genomic DNA Purification Kit Description |
designed for rapid and efficient purification of high quality genomic DNA from various mammalian cell culture and tissue sample, whole blood, bacteria and yeast; utilizes silica-based membrane technology in the form of a spin column |
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The GeneJET Genomic DNA Purification Kit Principle |
Samples are digested with Proteinase K in either the supplied Digestion or Lysis Solution. RNA is removed by treating the samples with RNase A. The lysate is mixed with ethanol and loaded on the purfication column where the DNA binds to the silica membrane. Impurities are removed by washing the column with prepared wash buffers |
