front 1 What are the cell dimensions of eukaryotic cells? | back 1 10-100 um |
front 2 What are the cell dimensions of intracellular organelles? | back 2 0.01-10um |
front 3 What are the main biomolecules? | back 3 proteins, rna, and dna |
front 4 What is the function and dimensions of proteins? | back 4 They are the molecular machines involved in all cell functions. 2-20nm |
front 5 What is the function and dimensions of dna? | back 5 They store information on the structure of all proteins. Diameter - 2nm |
front 6 What is the function of rna? | back 6 They transfer DNA to protein, protein synthesis, regulate of gene expression, and catalysis. |
front 7 What is the flow of genetic information (central dogma)? | back 7 DNA to mRNA to protein |
front 8 What is the largest intracellular organelle? | back 8 nucleus |
front 9 As particles move to the bottom of a tube during centrifugation, the centrifugal force acting on them... | back 9 increases |
front 10 What forces are important for particle sedimentation during centrifugation? | back 10 friction and buoyancy |
front 11 What happens to particles in solution during centrifugation? | back 11 They move towards and away from the rotation center. |
front 12 The friction force acting on a solid particle slowly moving in a liquid depends on... | back 12 particle shape, size, and the interaction between molecules of the liquid |
front 13 Sedimentation velocity of thin linear particles is... | back 13 not dependent on the particle length |
front 14 Sedimentation velocity of spherical particles is... | back 14 proportional to square of particle diameter |
front 15 As particles settle, will particle density be more or less than solution density? | back 15 Particle density will be greater. |
front 16 What is sedimentation velocities? | back 16 when particles are separated based on different velocities |
front 17 Sedimentation velocity can b increased by increasing what? | back 17 g (gravity) |
front 18 Friction coefficient f depends on what? | back 18 size and shape |
front 19 Denser particles of the same size and shape move... | back 19 faster |
front 20 What affects both volume and friction coefficent? | back 20 shape and size |
front 21 True or false. Sedimentation velocity is proportional to the ratio of particle volume and friction coefficient. | back 21 true |
front 22 Bigger spherical particles of equal density move what? | back 22 faster |
front 23 For sedimentation of biological particles, is the dependence on size more important than the dependence on density? | back 23 Yes |
front 24 Velocity-dependant seperation of near spherical biological particles is determined by what? | back 24 size differences |
front 25 Bigger biological particles have what? | back 25 larger sedimentation velocities |
front 26 The thin linear particle shape is common for what? | back 26 short double-stranded DNA, denatured RNA, denatured proteins, and cytoskeleton filaments. |
front 27 Can thin linear particles be separated by length? | back 27 no |
front 28 Linear thin particles of equal radius but different length have the same what? | back 28 sedimentation velocity |
front 29 what is the Svedberg coefficient? | back 29 the sedimentation velocity normalized by the centrifugal acceleration |
front 30 What does "s" depend on in the svedberg coefficient? | back 30 Particle properties. (ribosomes are measured in s unit) |
front 31 What is the organelle extraction by differential centrifugation? | back 31 there are several cycles run with increasing rotation speed and duration, after each run the largest particles will form a pellet at the bottom of the tube and it will be removed. |
front 32 Would particles with different sedimentation velocities
form | back 32 No |
front 33 What is the density gradient? | back 33 its when heavier fluid goes on top and will eventually sink due to gravity |
front 34 What is convection? | back 34 the flow and mixing of the suspension caused by density gradients in the centrifugation field. |
front 35 What is rate zonal centrifugation? | back 35 it is centrifugation in a density gradient. When a particle mixture is placed on top and a solution of sucrose is selected so that particle density is greater than solution density. |
front 36 What happens in centrifugation in a density gradient? | back 36 When particles move at a different speed, they form discrete bands, which are determined by sedimentation speed, which is dependent on particle size, density, and shape. |
front 37 Isopycnic centrifugation | back 37 When a particle floats or sinks to its equilibrium point where particle density equals the density of the solution. |
front 38 What does isopycnic mean? | back 38 same density |
front 39 Svedberg coefficient depends on what? | back 39 particle size, shape, and density |
front 40 The purpose of a density gradient in the Rate Zonal centrifugation is to what? | back 40 prevent convection |
front 41 The most important factor responsible for cell separation in Rate Zonal centrifugation is cell? | back 41 size |
front 42 Isopycnic centrifugation separates particles according to their differences in... | back 42 density |
front 43 Which forces determine electrophoresis of bio-molecules? | back 43 electrical and friction |
front 44 Electrophoretic velocity of short DNA molecules in water-based electrolyte is? | back 44 not dependant on molecule length |
front 45 Gel is solidified for DNA slab electrophoresis in order to... | back 45 prevent convection |
front 46 During electrophoresis, DNA molecules move towards what? | back 46 the anode |
front 47 During electrophoresis, native protein molecules move toward... | back 47 anode or cathode |
front 48 SDS-PAGE separates proteins based on differences in protein... | back 48 amino acid chain length |
front 49 Which molecules can be separated by electrophoresis in water-based electrolyte (without gel)? | back 49 non denatured RNA, non denatured proteins, and denatured proteins |
front 50 What is electrophoresis? | back 50 movement of electrically charged particles in a liquid medium caused by electric force |
front 51 If particles move with different velocities they do what? | back 51 they become spatially separated |
front 52 The separation factor ratio depends on what 3 properties? | back 52 1. charge 2. shape 3. size |
front 53 What is the source of DNA charge? | back 53 phosphate groups |
front 54 If DNA is negatively charged then the charge is what? | back 54 distributed uniformly |
front 55 How does the short double stranded DNA straighten out? | back 55 when the charged groups repulse one another |
front 56 what is the shape of short dsDNA? | back 56 linear or rod shaped |
front 57 Is separation of short dsDNAs of variable length in water-based electrolyte possible? | back 57 No |
front 58 EP of short dsDNA in a gel. | back 58 Double-stranded DNAs within a certain length range can be separated by EP in a gel based on their length differences |
front 59 What is the the gel? | back 59 A mesh of fibers forms pores filled with electrolytes through which DNA molecules can move. The gel can be liquid or solid. |
front 60 True or false. Longer molecules move through gel slower than shorter molecules | back 60 True |
front 61 What is slab gel EP? | back 61 when a slab of gel is solidified to prevent convection. This occurs when samples are loaded on the cathode side and negatively charged DNA move towards the anode. |
front 62 What is the field strength of gel? | back 62 5-20 V/cm |
front 63 Length measurements are done by what? | back 63 DNA ladders |
front 64 What is capillary gel EP? | back 64 It is run in thin capillaries so that gel can be liquid and require no convection and it is rapid dissipation of resistive heat. |
front 65 Under denaturing conditions, EP separation of ssDNAs or RNAs
is | back 65 their length |
front 66 What is the length speration range of EP gels? | back 66 function of gel pore size which depends on gel type and concentration. |
front 67 polyacrylamide gel | back 67 small pores, length less than 1000 bp, and high senstivity |
front 68 agarose gel | back 68 large pores, length less than 50,000 bps, and low sensitivity |
front 69 the net charge of a native protein can be what? | back 69 positive, negative, or either |
front 70 What is the most common method for electrophoretic protein separation? | back 70 SDS-PAGE (Sodium Dodecyl Sulfate-PolyAcrylamide Gel Electrophoresis) |
front 71 Steps of SDS-PAGE | back 71 1. denature proteins 2. treat with SDS which coats proteins with negative charges 3. repulses negative charges by straightening out proteins 4. treat proteins behave like short dsDNA |
front 72 SDS-treated proteins move toward the what? | back 72 anode |
front 73 True or false. Protein charge is proportional to the protein length | back 73 true |
front 74 SDS PAGE data presentation | back 74 Using a protein ladder and molecular weight |
front 75 The level of (de)protonation depends on what? | back 75 the solution pH |
front 76 When pH decreases and H increases what happens to protein charge? | back 76 it increases |
front 77 When pH increases and H decreases what happens to protein charge? | back 77 it decreases |
front 78 true or false. Each protein has its own unique pl value | back 78 true |
front 79 What is isoelectric focusing? | back 79 Separation of native proteins based on differences in their pl values |
front 80 A stable gradient of pH is created in what? | back 80 a gel |
front 81 Where is the anode located? | back 81 low pH - positive |
front 82 Where is the cathode located? | back 82 high pH - negative |
front 83 True or false. IEF is independent of protein MW | back 83 true |
front 84 When a molecule reaches its pl point what happens? | back 84 its charge becomes 0 and it stops |
front 85 2D EP happens in what 2 steps? | back 85 1. IEF - isoelectric focusing 2. SDS-PAGE |
front 86 Charge of which molecules is affected by changes of solution pH typical for intracellular environment? | back 86 Native proteins |
front 87 In the Isoelectric Focusing method, the cathode ( (-) electrode) is placed in the region with? | back 87 high pH |
front 88 The "housekeeping" RNAs are used in Northern blotting for... | back 88 measuring relative quantities of target RNAs |
front 89 The role of albumin in Western blotting is to... | back 89 block non-specific antibody binding |
front 90 The nitrocellulose membrane used in Western blotting binds... | back 90 all proteins |
front 91 Which protein property makes it a candidate for the role of a “housekeeping” protein in Western blotting? | back 91 stability of protein quantity |
front 92 How do we identify specific bio-molecule? | back 92 through staining with light/raditaion labels |
front 93 What are the DNA/RNA labels? | back 93 hybridiziation probes |
front 94 What are the protein labels? | back 94 immunistaining probes |
front 95 What are the 3 blotting methods? | back 95 1. Southern - DNA 2. Northern - RNAs 3. Western - proteins |
front 96 What are the blotting steps? | back 96 1. Gel electrophoresis |
front 97 what is the hybridization probe | back 97 a bio-conjugate of cDNA and reporter molecule |
front 98 Hybridization steps | back 98 1. target DNA/RNA and probe are denatured 2. The probe is added and binds to the target 3. light/radiation is detected |
front 99 Northern blotting | back 99 detects a specific mRNA in a sample gene expression |
front 100 What are the steps of northern blotting? | back 100 1. denature 2. separate by gel EP 3. Transfer to a membrane 4. Hybridization with probe 5. visualize |
front 101 Relative mRNA amounts from different samples are measured by comparing what? | back 101 band intensities |
front 102 true or false. mRNA is unstable | back 102 true |
front 103 Immunolabeling probe - Western blotting | back 103 a bio-conjugate of antibody and reporter molecule |
front 104 Western blotting steps | back 104 1. SDS PAGE 2. transfer proteins from gel to membrane 3. block antibody binding to membrane 4. label proteins with a primary antibody 5. label proteins with secondary antibody 6. detect the protein by optical imaging |
front 105 examples of housekeeping proteins | back 105 actin, tubulin, and GAPDH |
front 106 Genomics | back 106 analysis of organisms’ entire genomes or a large number of genes |
front 107 Functional genomics | back 107 global analysis of gene expression; identification and quantitative measurements of all (or many) mRNAs |
front 108 Proteomics | back 108 global analysis of cell protein content; identification and quantitative analysis of all (many) proteins |
front 109 What is chromatography | back 109 A family of techniques for rapid separation of complex mixtures
of |
front 110 2 phases of chromatography | back 110 1. stationary phase (gel, liquid, paper) 2. mobile phase (liquid or gas) |
front 111 Ion exchange chromatography | back 111 Stationary - resin beads with covalently bound charged molecules mobile - an electrolyte buffer with a certain ionic strength and pH |
front 112 WHat are the resin types in ion exchange chromatography? | back 112 anion exchanger- the resin has positive charges, it binds anions cation exchanger-the resin has negative charges, it binds cations |
front 113 What is elution? | back 113 step when main molecule separation takes place |
front 114 Ionic elution | back 114 Increase salt concentration and protein molecules are displaced by negative or positive ions. Separation is determined primarily by protein charge |
front 115 pH elution | back 115 Increases or decrease pH toward protein pl Separation is determined primarily by protein pl |
front 116 Molecular exclusion chromatography | back 116 Stationary phase- porous gel beads separation is based on molecule size |
front 117 In cation exchange chromatography, what happens to protein charge? | back 117 it decreases |
front 118 In anion exchange chromatography, what happens to protein charge? | back 118 it increases |
front 119 Partition chromatogrpahy | back 119 stationary phase - a thin liquid film formed on the surface of an inert solid support mobile phase - another liquid solvent |
front 120 Paper chromatography | back 120 stationary phase - bound h2O mobile phase- another solvent less polar than h2O (solvent moves up sheet using capillary forces) |
front 121 Affinity chromatogrpahy | back 121 stationary phase - contains ligands immobilized on beads that bind a targeted protein Separation is based on specific protein-ligand affinity |
front 122 Absorption chromatography | back 122 stationary phase - a solid adsorbent (silica gel (SiO2) or alumina (Al2O3) mobile phase - typically, liquid (e.g., ether, ethanol), or gas |