Resin-purified digoxigenin-labeled DNA probes: an efficient protocol for mapping and fingerprinting sugarcane.
Nonradioactive Southern blotting using digoxigenin (DIG) has become routinely applied to the analysis of single-copy genes for genetic mapping because it is fast and safe. Previous studies indicate that DIG-labeled probes are suitable for single-gene detection in less complex genomes, but their efficient application to mapping a large octoploid genome has not been discussed. We developed a stream-lined procedure for nonradioactive restriction fragment length polymorphism mapping and DNA fingerprinting of sugarcane that combines DIG-11-dUTP and anion-exchange chromatography. In this report, we show that anion-exchange chromatography provides a reliable and simple technique for the resin-purification of large numbers of DIG-labeled DNA fragments 0.3-3.0 kb in size, and it is essential in minimizing contaminants and nonspecific signal.
Ferrero
E A
EA
Hawaiian Sugar Planters' Association Experimental Station, Aiea, USA.
Fitch
M M
MM
Moore
P H
PH
Wu
K K
KK
D000837:Anion Exchange Resins / Q000737:chemistry*; D016477:Artifacts; D002852:Chromatography, Ion Exchange / Q000379:methods; D002874:Chromosome Mapping / Q000379:methods*; D016172:DNA Fingerprinting / Q000379:methods*; D015342:DNA Probes / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics; D004076:Digoxigenin; D012680:Sensitivity and Specificity
D000837:Anion Exchange Resins; D015342:DNA Probes; D018744:DNA, Plant; D004076:Digoxigenin