Mapping of Deletions and Substitutions in Heteroduplex DNA Molecules of Bacteriophage Lambda by Electron Microscopy

Abstract

Electron microscopy of heteroduplex DNA molecules, composed of one strand of Escherichia coli phage λ⁺ DNA annealed to the complementary DNA strand of a λ deletion or substitution mutant, permits visualization, as well as precise measurements and mapping, of the unpaired single-stranded regions of nonhomology in the otherwise double-stranded molecules. In the λb2 mutant, the central segment (13 percent) of the λ⁺ DNA molecule is shown to be deleted. In the hybrid phages λⁱ⁴³⁴ and λⁱ²¹ a segment of the right arm of the λ⁺ genome (5.5 or 7.6 to 9 percent) is replaced by the corresponding immunity regions of phage 434 (3.3 percent or phage 21 (4 percent) DNA. The b5 region in the λb5 mutant appears to be identical to the i²¹ segment. From these data it is possible to estimate the size and posiion of those λ genes which are replaced by the i⁴³⁴ and i²¹ segments. The method permits preparing complete physical maps of viral genomes with a precision heretofore unattainable.