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|a eng
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| 100 |
1 |
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|a Cho, Yong-Jig
|e verfasserin
|4 aut
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| 245 |
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|a Systematic analysis of intrinsic factors affecting differential display
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|c 2002
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| 336 |
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|a Text
|b txt
|2 rdacontent
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|a ohne Hilfsmittel zu benutzen
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|a Date Completed 13.11.2002
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|a Date Revised 28.09.2018
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|a published: Print
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|a Citation Status MEDLINE
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|a Differential display (DD) is a widely used method for identifying differentially expressed genes. To improve further the efficiency and reproducibility of the method, this report systematically examines four critical parameters of standard DD-PCR. Specifically, the study determined the optimal annealing temperature, elongation time, dNTP concentration, and arbitrary primer concentration. By using a thermal cycler that was capable of displaying a temperature gradient across a PCR plate, it was possible to determine (in a single experiment) the effect of different annealing temperatures. The optimal annealing temperaturefor a 13-mer arbitrary primer fell within a broad range of 40 degrees C-50 degrees C. Elongation times over a range of 30-120 s worked best. The optimal concentration for dNTPs was within a very broad range of 2-50 microM, with higher amounts allowing for greater pipetting accuracy. The most favorable concentration for the arbitrary primer was also within a broad range of 0.1-2.0 microM. A primer concentration below this range greatly reduced the efficiency of the amplification process. In conclusion, the experimental findings delineated the best possible DD conditions for a more reliable assessment of differential gene expression
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|a Research Support, U.S. Gov't, P.H.S.
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| 650 |
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|a Technical Report
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| 650 |
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|a Journal Article
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| 700 |
1 |
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|a Prezioso, Vincent R
|e verfasserin
|4 aut
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| 700 |
1 |
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|a Liang, Peng
|e verfasserin
|4 aut
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| 773 |
0 |
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|i Enthalten in
|t BioTechniques
|d 1993
|g 32(2002), 4 vom: 05. Apr., Seite 762-4, 766
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|x 0736-6205
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