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When restriction-active Escherichia coli cells (R+P1m+P1) are transformed with the pSF2124 plasmid, a common vector in experimental gene transfer, the efficiency of transformation (e.o.t.) is lowered by 2 orders of magnitude compared with restriction-negative (r-P1m-P1 or r-P1m+P1) recipient cells due to restriction of the pSF2124 DNA by endoR.EcoP1. Preinfection of r+P1m+P1 cells with UV-inactivated ocr+ phages (T3, T7, T3sam-) still able to express their early genes protects the plasmid DNA against restriction by endoR.EcoP1: The e.o.t. of r+P1m+P1 recipient cells with pSF2124 attains the same high value as that of r-m- cells. The specific role of the ocr+ gene function was demonstrated by the use of ocr- mutants (T3/R7, T7/D111): Preinfection with such phage mutants does not increase the e.o.t. of r+P1m+P1 cells. An unspecific e.o.t. alteration of restriction-negative (r-m-) recipient cells by ocr+ or ocr- phages was excluded. The ocr+ gene function can be exploited to protect pSF2124 against DNA restriction. The recipient cells survive the process of phage preinfection and transformation and stably replicate themselves as well as the plasmid DNA.

Transfection by DNA isolated from bacteriophage T3 was studied using Escherichia coli 921/0 as host. The following conditions were found optimal: Competent E. coli 921/0 were obtained by harvesting the bacteria at the onset of late exponential growth (5 X 10(8) cells/ml) and treating the latter with 0.05 M CaCl2. Hereafter, the microbes were suspended in 50 mM Tris-HCl buffer (pH 7.2) and the concentration adjusted to 7 X 10(9) cells/ml. T3 DNA was added and the suspension kept at 0 degrees C for 15 min. Determination of the number of infectious centers was then carried out in the usual way. The efficiency of transfection under these conditions amounted to 10(4) p. f. u./microgram DNA. Preincubation of competent bacteria with T4 DNA at 0 degrees C before the addition of T3 DNA reduced the number of infectious centers. However, if T3- and T4 DNA were added simultaneously no decrease of the transfection efficiency occurred. Calf thymus DNA was without influence on transfection.

In an enzyme-specific drug screening system nalidixic acid and 3'-FTdR, inhibitors of DNA synthesis, both reduce the growth of wild type and temperature-sensitive point mutants of phage T3 with different efficiencies. The wild type shows the strongest sensitivity against the drugs, while an exonuclease mutant is the most insensitive variant. The DNA polymerase mutants exhibit an intermediate degree of inhibition. The anthracycline antibiotics violamycin BI and adriblastin which preferentially inhibit RNA synthesis show the same degree of inhibition for all mutants. This is true also for the RNA synthesis inhibitor lambdamycin, which is identical with chartreusin. The protein synthesis inhibitors chloramphenicol and o-phenanthroline, a chelating agent, impair all mutants to the same extent. Our data confirm the hypothesis that structural variants of essential viral enzymes, when compared with the wild type should reveal different sensitivities against specific inhibitors and show that this T3 system could be used for the indication of specific inhibitors of DNA synthesis.

The closely related phages T3 and T7 exhibit different growth patterns on Escherichia coli W hosts cells (E. coli K12 derivatives). T7 grows normally while T3 does not adsorb. T3hw mutants displaying a T7-like host range were isolated and described.