Fast and sensitive silver staining of DNA in polyacrylamide gels.
The photochemically derived silver stain of nucleic acids in polyacrylamide gels originally described by Merril et al. (1981, Science 211, 1437-1438) was modified to reduce unspecific background staining and increase sensitivity (down to 1 pg/mm2 band cross-section). Detection limits for double-stranded DNA fragments from HaeIII endonuclease digests of phage phi X174 were maintained despite eliminating oxidation pretreatment of fixed gels and reducing silver nitrate concentration. Preexposure to formaldehyde during silver impregnation enhanced sensitivity and the inclusion of the silver-complexing agent sodium thiosulphate in the image developer decreased background staining. Higher formaldehyde concentration during image development resulted in darker bands with good contrast. The procedure almost halves the number of steps, solutions and experimental time required and can be used for the staining of DNA fragments in polyacrylamide gels bound to a polyester backing film by controlling temperature during image development. We have applied this improved staining procedure for the routine analysis of complex DNA profiles generated by DNA amplification fingerprinting (DAF).
Bassam
B J
BJ
Institute of Agriculture, University of Tennessee, Knoxville 37901-1071.
Caetano-Anollés
G
G
Gresshoff
P M
PM
D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D016172:DNA Fingerprinting / Q000379:methods*; D004279:DNA, Viral / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005557:Formaldehyde; D008969:Molecular Sequence Data; D011091:Polyesters; D012680:Sensitivity and Specificity; D012834:Silver; D012835:Silver Nitrate; D013194:Staining and Labeling / Q000379:methods*; D013696:Temperature; D013885:Thiosulfates
D004279:DNA, Viral; D011091:Polyesters; D013885:Thiosulfates; D005557:Formaldehyde; D012834:Silver; D004247:DNA; D012835:Silver Nitrate