Studies on the lambda resistant genes isolated from brevibacterium albidum and proteus vulgaris

Abstract

Genes, which make their host resistant to lambda, were isolated from $\underline{Brevibacterium}$ $\underline{albidum}$ ATCC15831 and Proteus vulgaris ATCC13315 and introduced into Escherichia coli HB101. The clones transformed by one of the recombinant plasmids, pRMG101 or pRMG216, were totally resistant to virulent lambda and N4, but sensitive to 080, T4, and T7. $\underline{E.}$ $\underline{coli}$ HB101 (pRMG101) was still resistant even when the cell was mixed with 30 times excess numbers of the phages, lambda and N4. Radioactively labelled plasmid DNAs, the pRMG101 and pRMG216 were hybridized to the chromosomal DNAs of $\underline{P.}$ $\underline{vulgaris}$ DNA and $\underline{B.}$ $\underline{albidum}$, respectively, indicating that the inserted foreign DNA fragments possessing lambda and N4 resistant activities in pRMG101 and pRMG216 were those bacterial organisms. There were no type II restriction activities found in the extracts of $\underline{E.}$ $\underline{coli}$ HB101 (pRMG101) or $\underline{B.}$ $\underline{coli}$ HB101(pRMG216) which were fractionated by heparinagarose column chromatography. While the two modification activities were detected from the extract, lambda DNA methylated by these modification enzyme were completely digested by each one of the endonucleases, Bal I, BamH I, Pvu I, Pvu II, and Xba I. Furthermore, a recombinant plasmid, pRMG101, was completely digested by Pvu I or Pvu II. Sequenced nucleotides revealed that the genes in pRMG101 contained one transcriptional unit with a representative promotor region and two open reading frames of the sizing, 500 and 960 base pairs (gene A and gene B, respectively). The gene A alone could not exhibited the resistance to the phages. In the case of the gene B, we were not able to subclone the gene B without the gene A to the downstream of any functional promotors. When phage DNAs were transfected into the clone carrying pRMG101AB (a subcloned plasmid) or pRMG216, not only 080 but also N4 a...