--Silvana Faccio Dolfini
Recent work (Miller et al., Science 209:395, 1983; Mezzanotte et al., Cytogenet. Cell Genet. 36:562, 1983) has proved that exposure of fixed chromosomes in cytological preparations to restriction endonucleases results in cleavage of DNA at specific base sequence targets.
The enzyme most frequently used is AluI, which selectively digests DNA (restriction sequence AGCT), producing a linear differentiation along a chromosome, consisting of alternating regions where DNA has been cleaved (and consequently extracted) and regions not affected by the enzyme (AluI-bands). In human, great ape, mouse and Drosophila chromosomes an overlapping between AluI-bands and localization of satellite DNAs has been demonstrated. Recent data from Vicia faba chromosomes (Frediani et al., TAG 75:46, 1987) favour the hypothesis that the induction of AluI-bands depends not only on the presence of a specific DNA base sequence, but also on a suitable configuration of chromatin.
In this note I report some preliminary attempts to induce AluI-bands on maize chromosomes from two inbred lines (H84 and W117). Permanent root tip preparations were digested by placing enzyme solution (15 units of AluI in 100ul of the appropriate buffer) over the slides, covering with a coverslip and incubating overnight at 37 C. The chromosomes were then stained with 3% Giemsa.
AluI treatment on H84 chromosomes induces discrete bands on precise sites of metaphase (Fig.1c) and prophase (Fig.1b) chromosomes and intensely stained dots on interphase nuclei (Fig.1a). Different results in terms of number, size and position of the bands are observed on W117 chromosomes.
Fig. 1. AluI digestion of interphase nuclei (a, x2000), prophase (b, x1600) and metaphase (c, x2400) chromosomes from H84 inbred line.
These preliminary observations open the way to a series of investigations on the homology between the bands revealed by AluI treatment and the heterochromatic knobs visible in pachytene and the relationships between AluI-bands and knobs distinguishable by C-banding in mitotic chromosomes.
Finally, maize chromosomes, characterized by the presence of different
classes of heterochromatin (centromeric, knob, B-chromosomes, NOR), may
help to elucidate the mechanism of AluI-bands appearance.
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