Transport of Donor Deoxyribonucleic Acid into the Cell Interior of Thymine-starved Bacillus subtilis with Chromosomes Arrested at the Terminus

Abstract

The chromosomes of a tryptophan − , thymine − double auxotroph of Bacillus subtilis were uniformly aligned at the chromosome terminus by an amino acid starvation treatment. By subsequent incubations, the starved culture was rendered competent, while its state of synchronous chromosome arrest was maintained by thymine starvation. The competent, chromosome-arrested cells were transformed for three unlinked markers, located in two different chromosome regions. Shortly after addition of deoxyribonucleic acid, the cell walls were removed with lysozyme in a medium containing deoxyribonuclease and no thymine, and the protoplasted culture was assayed for single and double transformants. It was found that markers both near and distant from the terminus entered freely into the cell interior. There was no important difference in the relative frequency of entry of different markers between synchronously arrested cells and nonsynchronized control cultures. It is concluded that entry of a given marker into the cell interior can occur even if the replication site of the chromosome is stationary at a location distant from the locus of the resident homolog of the entering marker. A mechanism of donor deoxyribonucleic acid entry involving homology at the replication fork is excluded.