Detection of single-copy genes in DNA from transgenic plants by nonradioactive Southern blot analysis
- 1 February 1997
- journal article
- Published by Springer Nature in Molecular Biotechnology
- Vol. 7 (1) , 79-84
- https://doi.org/10.1007/bf02821545
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.Keywords
This publication has 14 references indexed in Scilit:
- Quantitation of PCR products with chemiluminescence.1995
- Genotype-independent transformation of lettuce usingAgrobacterium tumefaciensJournal of Experimental Botany, 1994
- Nonradioactive labeling of oligonucleotides in vitro with the hapten digoxigenin by tailing with terminal transferaseAnalytical Biochemistry, 1991
- Nonradioactive labeling of probe with digoxigenin by polymerase chain reactionAnalytical Biochemistry, 1990
- Non-radioactive labeling of RNA transcriptsin vitrowith the hapten digoxigenin (DIG); hybridization and ELISA-based detectionNucleic Acids Research, 1990
- Non-radioactive Labeling and Detection of Nucleic Acids. II. Optimization of the Digoxigenin SystemBiological Chemistry Hoppe-Seyler, 1990
- 1,2‐Dioxetanes: Novel chemiluminescent enzyme substrates. Applications to immunoassaysJournal of Bioluminescence and Chemiluminescence, 1989
- Stable transformation of maize after gene transfer by electroporationNature, 1986
- Enzymatic Amplification of β-Globin Genomic Sequences and Restriction Site Analysis for Diagnosis of Sickle Cell AnemiaScience, 1985
- A technique for radiolabeling DNA restriction endonuclease fragments to high specific activityAnalytical Biochemistry, 1983