This set of Gene Manipulation Interview Questions and Answers for Experienced people focuses on “Basic Laboratory Techniques – 7”.
1. An ideal oligonucleotide sequence should remain hybridized when it is _____ percent homologous to the target.
Explanation: The availability of the exact sequence of oligonucleotides allows conditions for hybridization and stringency washing to be tightly controlled so that the probe remains hybridized when it is 100% homologous to the target.
2. The “Wallace Rule” is used to determine which of the following?
a) Melting temperature
Explanation: Stringency is commonly controlled by adjusting the temperature of the wash buffer. The “Wallace Rule” is used to determine the appropriate stringency wash temperature.
3. Which temperature is measured in the Wallace rule?
Explanation: The Wallace rule makes use of wash temperature to determine appropriate stringency conditions. It is usually a preliminary step.
4. In filter hybridization with oligonucleotide probes, the hybridization is performed _____ degrees below TM.
Explanation: In filter hybridization with oligonucleotide probes, the hybridization is performed at a lower temperature than the actual melting temperature.
5. For every mismatched base pair, a further 5⁰C reduction is necessary.
Explanation: In filter hybridizations with oligonucleotide probes, the hybridization step is usually performed at 5⁰C below TM for perfectly matched sequences.
6. The design of oligonucleotide is critical for ____________
d) Temperature control
Explanation: The design of oligonucleotides for hybridization experiments is critical to maximize hybridization specificity.
7. Which of the following greatly influences the stability of the resultant hybrid?
a) AT content
b) GC content
c) Oligonucleotide weight
d) Melting temperature
Explanation: The GC content of oligonucleotide influences the stability of the resultant hybrid and hence the determination of appropriate stringency conditions.
8. Autoradiography involves production of _____________
Explanation: The localization and recording of a radiolabel within a solid specimen is known as autoradiography and involves the production of an image in a photographic emulsion.
9. Emulsions in autoradiography, contain a clear phase of ____________
Explanation: Emulsions consist of silver halide crystals suspended in a clear phase composed mainly of gelatin. Silver ions are later converted into silver atoms.
10. Which of the following will convert silver ion to a silver atom in the emulsion?
a) Alpha particle
d) Beta particle
Explanation: When a beta-particle or gamma-ray from a radionuclide passes through the emulsion, the silver ions are converted to silver atoms.
11. The radioactive emissions produce which color on the developed autoradiograph?
Explanation: In direct autoradiography, the sample is placed in intimate contact with the film and radioactive emissions produce black areas on the developed autoradiograph.
12. Direct autoradiography is not suited for particles such as phosphorus.
Explanation: Direct autoradiography has not suited the detection of highly energetic beta-particles, such a phosphorus-32 or gamma-rays.
13. How is emitted energy converted to light in Autoradiography?
Explanation: Indirect autoradiography describes the technique by which emitted energy is converted to light by means of a scintillator, using fluorography.
14. Which of the following is used to improve the detection of weak beta-emitters?
Explanation: Fluorography is used for the detection of weak beta-emitters. Analysis of such compounds is necessary and hence unique techniques are used.
15. Intensifying screens are sheets of solid ____________ scintillator.
Explanation: Intensifying screens are sheets of a solid inorganic scintillator which are placed behind the film. Emissions are absorbed by the film.
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