The structural organization of mouse metaphase chromosomes

The binding of highly purified anti-nucleoside antibodies to mouse (Mus musculus) metaphase chromosomes was studied by an immunofluorescence technique. The chromosomal DNA was denatured by one of two selective denaturation procedures because these antibodies reacted with single stranded but not native DNA. After ultraviolet irradiation (UV), which produced single stranded regions primarily in AT rich DNA, the binding of antiadenosine (anti-A) produced a pattern of fluorescent bands similar to that produced by quinacrine (Q-bands). Additional foci of bright fluorescence were observed at the centrometric (C-band) regions, which are known to contain AT rich satellite DNA. After photooxidation, which produced single stranded regions in GC rich DNA, the binding of anti-A produced a fluorescent banding pattern similar to the R-banding pattern seen after thermal denaturation and staining with coriphosphine O. After photooxidation, R-band patterns were also obtained with anti-cytidine (anti-C) and anti-5-methylcytidine (anti-M). After either UV irradiation or photooxidation, anti-M, but not anti-C, showed intense binding to the C-band regions of mouse chromosomes. — These findings led to the following conclusions: (1) Antibody banding patterns reflect the presence of a class of AT rich, GC poor DNA in chromosome regions which show bright quinacrine fluorescence and in the regions that contain the AT rich satellite DNA. (2) The alternate, quinacrine dull regions contain a relatively GC rich class of DNA which appears to be more highly methylated than the AT rich DNA in the Q-bright bands, but not the AT rich satellite DNA in the Q-dull C-bands. (3) 5-Methylcytosine residues occur in a sequence of mouse satellite DNA that contains both adjacent pyrimidines and guanine residues. The basic repeating unit of mouse satellite DNA is known to contain the sequence 5-GAAAAATGA-3 (Biro et al., 1975). Therefore, assuming the antibodies used could detect single bases in denatured DNA, the methylated sequence in mouse satellite DNA