DNA sequence mapping using electron microscopy.
DNA sequences can be mapped on chromosomes at high resolution in the electron microscope after hybridization with a nonisotopically labeled probe followed by detection with a two-step antibody reaction employing a colloidal gold tag. Hybridization probes can be modified with biotin-dUTP, digoxigenin-dUTP, dinitrophenyl-dUTP, or N-acetoxy-2-acetylaminofluorene (AAF). The availability of different sizes of colloidal gold particles permits the simultaneous detection of several sequences. In addition, low signals can be amplified either with an antibody sandwich scheme or by silver intensification. This technology is applicable both to TEM and SEM preparations of chromosomes, and we have used it to map a number of highly and moderately repeated sequences on whole mount metaphase chromosomes.
Narayanswami
S
S
Department of Molecular Biology and Biochemistry, University of California, Irvine 92717.
Hamkalo
B A
BA
D000099:Acetoxyacetylaminofluorene; D000818:Animals; D001483:Base Sequence; D001710:Biotin; D002874:Chromosome Mapping / Q000379:methods*; D004247:DNA / Q000737:chemistry*; D004076:Digoxigenin; D004140:Dinitrophenols; D006801:Humans; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron; D015336:Molecular Probe Techniques; D009693:Nucleic Acid Hybridization; D012091:Repetitive Sequences, Nucleic Acid
D004140:Dinitrophenols; D000099:Acetoxyacetylaminofluorene; D001710:Biotin; D004247:DNA; D004076:Digoxigenin