Effect of formaldehyde on the efficiency of hybridization of DNA immobilized on nitrocellulose filters.
We report in this study that under certain conditions formaldehyde interacts with DNA and makes it more efficient for hybridization on nitrocellulose filters. Hybridization signals of formaldehyde-treated DNA are stronger (up to 10 fold) as compared with that of the heat- or alkali-denatured DNA. Various parameters of the DNA-formaldehyde reaction are optimized as follows: (a) 6 x SSC, 10% formaldehyde, 60 degrees C, 20-30 min, reaction volume 10-200 microliters or (b) 6 x SSC, 5% formaldehyde, 98 degrees C, 15 min, reaction volume 10-200 microliters. Treatment of agarose gels after electrophoresis with formaldehyde improved both the transfer of DNA and the efficiency of hybridization. The following conditions are recommended for gel treatment: denaturation in 0.3 N NaOH, 1 M NaCl followed by neutralization with 0.5 M phosphate buffer, pH 7.0, containing 10% formaldehyde at 60 degrees C for 20 min.
Ivanov
I G
IG
Department of Botany, University of Toronto, Ontario, Canada.
AbouHaidar
M G
MG
D003101:Collodion; D004247:DNA / Q000032:analysis* / Q000235:genetics; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005557:Formaldehyde; D007202:Indicators and Reagents; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization / Q000379:methods*; D010761:Phosphorus Radioisotopes; D010957:Plasmids
D007202:Indicators and Reagents; D010761:Phosphorus Radioisotopes; D005557:Formaldehyde; D003101:Collodion; D004247:DNA