Detection of single-copy genes in DNA from transgenic plants by nonradioactive Southern blot analysis |
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Authors: | Matthew S McCabe J Brian Power Ad M M de Laat Michael R Davey |
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Institution: | (1) Plant Genetic Manipulation Group, Department of Life Science, University of Nottingham, University Park, NG7 2RD Nottingham, UK |
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Abstract: | Improvements to the sensitivity, speed, and reproducibility of digoxigenin (DIG)-labeled probes and chemiluminescent substrates
makes these compounds increasingly popular to detect nucleic acids. High sensitivity and low background are essential in Southern
blot analysis, particularly with plant DNA. This article describes a nonradioactive system to detect single-copy genes in
transgenic plants. Labeling using the polymerase chain reaction (PCR) was employed to produce highly sensitive and reusable
DIG-labeled probes. The background was reduced by immobilizing the DNA onto nylon filters by alkaline transfer and by minimized
gel handling; the signal-to-noise ratio was improved by modification of the detection procedure. |
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Keywords: | Digoxigenin chemiluminescent detection lettuce DNA |
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