Figure 2.

The effect of pH on Hsmar1 transposition. (A) Hsmar1 transposition reactions were carried out at the indicated pH values. A photograph of an ethidium bromide-stained agarose gel is shown. Inter., intermolecular transposition into an unreacted (circular) donor plasmid; Inter.*, the linear product of intermolecular transposition into the linear form of the donor plasmid; the IC is the simplest form of intramolecular integration product as illustrated in Figure 1A; Intra., topologically complex intramolecular insertion products also illustrated in Figure 1A. The plasmid backbone is an end product of the reaction and is used to quantify transposon excision from the donor. The circular ‘Inter.’ intermolecular product was identified after purification from the gel. Individual clones of these molecules were recovered by transformation in E. coli, and the size and structure of a representative selection were analyzed by restriction mapping and nucleotide sequencing. The linear topology of the ‘Inter.*’ product was assigned from the gel in Figure 3. Its position in the present gel is consistent with the size expected for a transposon insertion into a linear donor plasmid. The ‘Intra.’ intramolecular products were identified in Figure 3 (see text for details). (B) The in vitro hop assay for intermolecular transposition was performed at the indicated pH values.