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In this paper, we report the construction in vitro of chimerae between lambdoid replacement vectors (Murray et al., 1977) and the miniF Apr plasmid: pSC138 (Timmis et al., 1975). λF recombinants were shown to be chimerae between the λ and the F replicons. By genetical tests, we have demonstrated that both λ and F replication mechanisms are functional: the λF recombinant behaves as a non defective plaque forming phage on λ sensitive bacteria and establishes itself as a stable plasmid on recA F- homoimmune bacteria. In the extra-chromosomal state, the λF recombinant apparently retains the controlled autonomous replication and the FI incompatibility characteristics of the F plasmid. The potential experimental uses of these phages are discussed.

A chimeric β-lactamase encoding plasmid, containing the 4.4 Mdal β-lactamase plasmid and the 2.6 Mdal cryptic plasmid of Neiseria gonorrhoeae has been characterized by physical and biological methods. Digestion with restriction enzymes indicates the presence of the following restriction sites: 1 site: AccI, AvaI, HgiAI, HincII, MstI, PstI, PvuII, XbaI and XorI; 2 sites: HindIII and BamHI; 3 sites: BclI, Sau961 and AvaII; 6 sites: HinfI; >8 sites: AluI, BbvI, DdeI, HhaI, HpaII, MspI, Sau3A, TacI and TaqI. No restriction sites were found for the following: BglI, BglII, BstEII, EcoRI, EcoRII, HpaI, HphII, KpnI, SacI, SalI, SmaI, SstI, SstII, and XhoI. Five plasmid specific proteins have been identified by DNA directed in vitro protein synthesis (43K, 41K, 30K, 16K and 14K). The location on the physical map of the coding regions for each of these proteins has been determined by the following methods: using plasmid DNA restricted by various enzymes in an in vitro protein synthesis system; identifying promoter-containing regions by digesting plasmid DNA with DdeI, adding RNA polymerase and then determining which fragments are retained by nitrocellulose. This plasmid contains both parental phenotypes in that it encodes penicillin resistance and possesses the sequence necessary for uptake in the gonococcus. Transformation data indicate that this plasmid can function in both E. coli and N. gonorrhoeae and that growth in E. coli has no effect on the plasmid's ability to transform the gonococcus.

The lethal and mutagenic effects of 5 μg/ml N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) were maximal during the nuclear S-period of synchronously grownChlamydomonas reinhardtii. This was revealed by a 50% drop in survival and a 50- to 100-fold increase in the recovery of slow-growth mutants (up to 40% of the survivors) which were first recognized as small colonies on agar medium. Partial characterization of these isolates revealed about 50% to be stable on subculture, and several were demonstrated to be either acetate-dependent, dark-lethal (light-dependent), or acetate-sensitive mutants. There was no significant increase of lethality or of slow-growth mutants correlated with treatment during the chloroplast DNA replication phase of the cell-cycle. The results of genetic analysis with 13 mutants induced during the nuclear S-period were consistent with their nuclear origin. These analyses were hampered by the high proportion of lethality among the progeny of most crosses. It is concluded that the enhanced mutant induction among nuclear S-phase cells may indicate preferential mutagenesis of replication fork DNA and induction of multiple-closely-linked mutations, as in some bacteria. Consequently, forC. reinhardtii, caution should be exercised in drawing relationships between abnormal behavioral and biochemical phenotypes in MNNG-induced mutants.

The sedimentation coefficients of the NADPH: cytochrome-c oxidoreductase enzymes from wild-type and mutant strains of Aspergillus nidulans have been estimated by sucrose density gradient centrifugation. In the wild-type, two species of cytochrome-c reductase were found, with sedimentation coefficients of 13.7s and 7.6s respectively. The 13.7s species did not appear to be associated with the enzymes of nitrate reduction, whereas the 7.6s species was closely associated with NADPH: nitrate oxidoreductase. In mutant strains lacking nitrate reductase, a thir species of cytochrome-c reductase with a sedimentation coefficient of 4.5s was found. There is some evidence that this 4.5s cytochrome-c reductase is a subunit or breakdown product of nitrate reductase and a model is presented for the role of this 4.5s cytocnorome-c reductase in the assembly of the intact nitrate reductase molecule.

Mutations in the forked (f) gene of Drosophila cause deformation of bristles and hairs. Our molecular analysis showed the f gene to span more than 30 kb, and to encode two major RNAs, 6.0 and 2.5 kb long, both of which are prematurely terminated in gypsy and springer insertion mutants. These truncated RNAs were polyadenylated using putative polyadenylation signals within the 5′-LTR of the inserted retrotransposon. No evidence was found for effects of the retrotransposon insertions on the promoters for transcription of the 6.0 and 2.5 kb RNAs. In f 1 and f x , a single gypsy element was found to be inserted at identical sites in the second intron of region encoding the 2.5 kb f RNA and both truncated and wild-type sized RNAs were detected. Recessive mutations at suppressor of forked (su(f)) increased the fraction of wild-type sized RNAs considerably, suggesting that the wild-type su(f) product either stimulates premature termination at the gypsy LTR or inhibits normal splicing. In f 36a , a springer element inserted in the third exon of the region encoding the 2.5 kb f RNA completely suppressed the formation of apparently wild-type transcripts.

We examined the possibility that the ssb-1 and ssb-113 mutants exert some of their effects by interfering with the normal function of wild-type RecF protein. Consistent with this possibility, we found that recA803, which partially suppresses recF mutations, also partially suppresses both ssb mutations, as detected by an increase in UV resistance. No evidence was obtained for suppression of the defect in lexA regulon inducibility caused by the ssb mutations. Consequently we suggest that suppression occurs by increasing recombinational repair. In vitro tests of Ssb mutant and wild-type proteins revealed that the single-stranded DNA dependent ATPase activity of RecA protein is more susceptible to inhibition than the joint-molecule-forming activity. All three Ssb proteins inhibit the ATPase activity of RecA wild-type protein almost completely while under similar conditions they inhibit the joint-molecule-forming activity only slightly. Both activities of RecA803 protein were found to be less inhibited by the three Ssb proteins than those of RecA wild-type protein. This is consistent with the suppressing ability of recA803. We found no evidence to contradict the previously proposed hypothesis that ssb-1 affects recombinational repair by acting as a weaker form of Ssb protein. We found, however, only very weak evidence that Ssb-113 protein interferes directly with recombinational repair so that the possibility that it interferes with a normal function of RecF protein must remain open.