Organization of heterologous DNA inserts on the mouse meiotic chromosome core

Abstract

With simultaneous immunofluorescence and fluorescent in situ hybridization, we have determined the organization of native and heterologous DNA sequences relative to the cores of meiotic prophase chromosomes. The normal chromatin organization is demonstrated with probes of mouse sequences: a cosmid probe that identifies unique sequences and a 720 kb yeast artificial chromosome (YAC) probe that recognizes a specific region of the chromatin domain. The heterologous DNA consists of a 1.8 Mb insertion of 40 tandem head-to-tail phage λ LIZ vectors and of 11.4 Mb of bacterial/mouse DNA repeats. The lengthy λ insert is unusual in that it is not contained in the chromatin domain of chromosome 4 and in that it fails to form direct attachments to the chromosome core. The ends are attached indirectly, probably by means of the flanking mouse sequences. At late stages of meiotic prophase, while the terminal attachments remain the same, the λ DNA becomes highly compacted. Apparently, higher order condensation and core attachment are independent processes. The condensed inserts relax precociously at metaphase I. In the mouse heterozygous for the insert, the two sister inserts are usually merged, as are all four inserts in the homozygous mouse. Evidently chromatin loops with identical sequences can become associated during meiotic prophase. Mouse sequences within a heterologous DNA insert (repeats of bacterial plasmid pBR322 with a mouse β-globin insert) were observed to restore some degree of core attachment.