Acid depurination after field inversion agarose gel electrophoresis reduces transfer of large DNA molecules.
Field-inversion gel electrophoresis (FIGE) is an economical method for the resolution of DNA fragments 20 to 1000 kilobase pairs (kbp) in length, sizes beyond the resolving capabilities of normal agarose gel electrophoresis. A theoretical limitation to FIGE is the subsequent transfer of large DNA molecules to membrane supports for hybridization. After normal electrophoresis, uniform and rapid capillary transfer of DNA fragments from agarose gels has been previously reported to be facilitated by brief depurination of DNA with 0.25 N HCl. However, after FIGE we have found that brief treatment of large (greater than 200 kbp) linear DNA fragments with 0.25 N HCl reduces the extent of subsequent transfer by capillary methods. After FIGE and transfer, ethidium bromide staining of DNA suggests that acid treatment causes trapping of DNA within the agarose matrix.
Van Devanter
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Division of Medicine/Oncology, University of Texas Health Science Center San Antonio 78284-7884.
Von Hoff
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D004247:DNA / Q000302:isolation & purification*; D004270:DNA, Circular / Q000302:isolation & purification; D004271:DNA, Fungal / Q000302:isolation & purification; D004272:DNA, Mitochondrial / Q000302:isolation & purification; D004279:DNA, Viral / Q000302:isolation & purification; D003720:Densitometry; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004996:Ethidium; D004854:Herpesvirus 4, Human / Q000235:genetics; D009693:Nucleic Acid Hybridization; D011687:Purines / Q000302:isolation & purification*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013194:Staining and Labeling
D004270:DNA, Circular; D004271:DNA, Fungal; D004272:DNA, Mitochondrial; D004279:DNA, Viral; D011687:Purines; D004247:DNA; D004996:Ethidium