Genotyping of DNA using sequence-specific methyltransferases followed by immunochemical detection.
Modern molecular genetics relies on the ability to map the positions of genes on chromosomes, relative to known DNA markers. The first such DNA markers described were Restriction Fragment Length Polymorphisms, but any restriction endonuclease used for RFLP mapping is just one member of a restriction-modification pair. For each restriction endonuclease, there is a companion methyltransferase (MTase) that has the same DNA sequence specificity. Therefore, in principle, it should be possible to use MTases rather than restriction enzymes to detect polymorphic sites in DNA. We have used sequence-specific DNA MTases to detect polym orphisms in closely related viral pathogens. If at least one MTase recognition site is present in PCR-amplified DNA, then methyl groups are incorporated; if no MTase site is present, then methyl groups are not incorporated. When several different sequence-specific DNA MTase reactions are carried out, the pattern of methyl incorporation defines a DNA MTase genotype. DNA MTase Genotyping (DMG) can be used to rapidly diagnose heritable or infectious diseases, to immunochemically detect DNA at defined 2 to 8 base pair sites, or to characterize the amplicons by constructing ordered maps.
López
Osvaldo J
OJ
University of Buenos Aires and CONICET, Buenos Aires, Argentina.
Quintanar
André
A
Padhye
Nisha V
NV
Nelson
Michael
M
D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D016172:DNA Fingerprinting / Q000379:methods*; D019175:DNA Methylation; D005838:Genotype; D005914:Globins / Q000235:genetics; D018259:Herpesvirus 1, Human / Q000235:genetics; D018258:Herpesvirus 2, Human / Q000235:genetics; D007120:Immunochemistry; D008780:Methyltransferases; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011874:Radiometry; D019809:Streptavidin
D005914:Globins; D004247:DNA; D019809:Streptavidin; D008780:Methyltransferases