12) Which of the following statements is true about an electrophoretic mobility shift assay? A. Protein binding to DNA causes the DNA to migrate through the gel in an electric field faster than the DNA alone. B. The same binding affinity will be observed if the assay is run in 100 mM NaCl or 2 M NaCl. C. The gel consists of a pH gradient, such that the protein stops migrating when it reaches its isoelectric point. D. To observe the DNA-protein bound complex, the researcher should add SDS to ensure the protein has a negative charge. E. None of these answers are true.
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A. Protein binding to DNA increases the size of the DNA-protein complex, which slows down its migration through the gel. This statement is false. B. The binding affinity is affected by the salt concentration. Higher salt concentrations reduce the binding affinity. Show more…
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Which of the following statements is not correct? A) RNA molecules are negatively charged, therefore they migrate toward the anode (positive electrode) in a polyacrylamide gel placed in an electric field. B) DNA molecules are negatively charged, therefore they migrate toward the anode (positive electrode) in an agarose gel placed in an electric field. C) Negatively charged proteins migrate toward the anode (positive electrode) in a SDS-PAGE gel placed in an electric field. D) All proteins, regardless of their intrinsic charge properties, migrate toward the anode (positive electrode) in a SDS-PAGE gel placed in an electric field. E) Positively charged proteins migrate toward the cathode (negative electrode) in a SDS-PAGE gel placed in an electric field.
Dinesh S.
When DNA is subjected to electrophoresis in an agarose gel, shorter molecules migrate faster than longer ones. Closed-circular DNAs of the same size but different linking number also can be separated on an agarose gel: topoisomers that are more supercoiled, and thus more condensed, migrate faster through the gel-from top to bottom in the two gels shown below. A dye, chloroquine, was added to these gels. Chloroquine intercalates between base pairs and stabilizes a more underwound DNA structure. When the dye binds to a relaxed closed-circular DNA, the DNA is underwound where the dye binds, and unbound regions take on positive supercoils to compensate. In the experiment shown here, topoisomerases were used to make preparations of the same DNA circle with different superhelical densities $(\sigma) .$ Completely relaxed DNA migrated to the position labeled $\mathrm{N}$ (for nicked), and highly supercoiled DNA (above the limit where individual topoisomers can be distinguished) to the position labeled X. (FIGURES CAN'T COPY) (a) In gel $A$, why does the $\sigma=0$ lane (i.e., DNA prepared so that $\sigma=0,$ on average) have multiple bands? (b) In gel $\mathrm{B}$, is the DNA from the $\sigma=0$ preparation positively or negatively supercoiled in the presence of the intercalating dye? (c) In both gels, the $\sigma=-0.115$ lane has two bands, one a highly supercoiled DNA and one relaxed. Propose a reason for the presence of relaxed DNA in these lanes (and others). (d) The native DNA (leftmost lane in each gel) is the same DNA circle isolated from bacterial cells and untreated. What is the approximate superhelical density of this native DNA?
MCQ. Choose the correct answer. 1. Which of the following is not true for SDS in denaturing gel electrophoresis? a) It linearizes proteins b) It imparts negative charges to the linearized proteins c) It dissociates the subunit structures of a protein d) It breaks the disulfide bond and dissociates subunits 2. Electrophoretic mobility of a protein in a gel depends upon several factors. Which one of the following is inversely affecting the mobility of a protein? a) The charge of a protein b) The shape of a protein c) The viscosity of the medium d) Electric field applied 3. In isoelectric focusing, proteins are separated on the basis of their a) the relative content of positively charged residue only b) the relative content of negatively charged residue only c) the relative content of positively and negatively charged residues d) all of the above 4. How does Native PAGE differ from SDS-PAGE? a) No denaturing and reducing agents are used in a native gel. b) Only no denaturing agent used in native gel c) Only no reducing agent used in the native gel d) All of the above 5. SDS-PAGE is a discontinuous gel electrophoresis because it utilizes a) Different percentage of gels b) Different pH of the buffers c) Different composition of buffers d) All of the above e) None of the above
Suman K.
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