[Molecular cloning and functional analysis of DNA regions of plasmid RP4 determining the incompatibility properties].

Sau3A-generated DNA fragments determining incompatibility functions of the plasmid RP4 were cloned on the vectors pTK16 and pBR322. Inc+ recombinant plasmids were divided into two types: 1) expressing incompatibility only towards the homologous RP4 replicon, 2) expressing incompatibility - both towards the homologous RP4 replicon and towards the heterologous replicons of plasmids R906 and R751. For one member of the first type plasmids it was shown that the cloned Inc+-specific insertion derived from the region of location of the EcoRI restriction site.

[Properties of potential vectors--derivatives of the broad-host--range plasmid RP4].

Nonconjugative deletion and recombinant derivatives of the RP4 plasmid are constructed. The plasmids can be used as vectors because they have relatively small molecular weights, unique cleavage sites for enzymes EcoRI, XhoI, BamHI, PstI, KpnI, BglII, SalGI and HindIII (the plasmids pRP401 and pRP417 having six of these sites), and easily tested phenotypes (Tcr, Apr and Gal+). In addition, all of them retain the broad host range property.

[Molecular genetic studies of the DNA promotor regions in Bacillus thuringiensis subsp. galleriae 69-6. II. Increased level of gene expression resulting from IS1 element integration].

The 1.45 kb promoter containing HindIII fragment of Bacillus thuringiensis DNA promotes the expression of the tet gene of recombinant pPBT9 plasmid in Escherichia coli cells. Spontaneous mutants of this plasmid were isolated and analysed.

[Bacteriocinogenic plasmids of Bacillus thuringiensis].

When strains of Bacillus thuringiensis v. morrisoni or v. darmstadiensis were plated on solid medium, the appearance of oligosporogenic (Ospo) mutants and bacteriocin non-producing clones (Thc-) was observed.

[Homology of plasmids with a wide range of hosts].

According to blotting hybridization and heteroduplex analysis, plasmids R751, R906 and RP4 of Inc Pi group have continuous regions of homology. These homologous regions were mapped on the R751 and RP4-derived pRP401 deletion mutant DNAs. The plasmid pRP401 (m.

[Enterobacterial gene expression of antibiotic resistance under the control of Bacillus thuringiensis regulatory signals in gram-negative and gram-positive bacteria].

Two recombinant plasmids pPBT9 and pPBT74 carrying HindIII promoter-active DNA fragments of Bacillus thuringiensis onto the vector pGA24 were studied. The cloned fragment of pPBT9 plasmid has many homologous regions in Bac. thuringiensis genome.

[Comparative analysis of the P-1-group plasmids of incompatibility].

Wide host range plasmids (IncP-1) R906, R751 and R702 have several cleavage sites for BamHI, HindIII and EcoRI enzymes, in contrast to RP4 plasmid. Using these enzymes, deletion mutants of R906 plasmid have been obtained in vitro which only lost short DNA fragments (1 to 14 kb). A narrow host range pAV1 plasmid of the same incompatibility group has been transformed into the cells of Escherichia coli.

[Distribution, homology and cloning of cryptic Bacillus thuringiensis plasmids].

The 69-6 strain of Bacillus thuringiensis subsp. galleriae harbours at least 7 cryptic plasmids (pBTG1 - pBTG7) with molecular lengths 8,4 to 15,7 kb. According to hybridization analysis, the plasmid pBTG2 (8,7 kb) and other plasmids of the same host strain as well as cryptic plasmids of the strains belonging to 10 other serotypes of Bac.

Transformation of Bacillus thuringiensis subsp. galleria protoplasts by plasmid pBC16.

Protoplasts of the entomopathogenic bacterium Bacillus thuringiensis subsp. galleria were transformed by plasmid pBC16. The frequency of transformation was much lower than that of Bacillus subtilis.

[Cloning and the expression of the DNA promotor fragments of Bacillus thuringiensis in Escherichia coli cells].

The promoter-containing fragments of Bacillus thuringiensis subsp. galleria 69-6 DNA have been cloned on the pGA24 vector in Escherichia coli cells. The recombinant plasmids make cells resistant to tetracycline in a wide range.

Expression of E. coli genes carried by hybrids of plasmid RP4.

Hybrids of the RP4 plasmid, containing bacteriophage Mu and chromosomal genes of Escherichia coli, were transferred into Salmonella typhimurium, Pseudomonas putida, Pseudomonas aeruginosa and Proteus mirabilis. The individual genes of the arginine, histidine, leucine and threonine operons were expressed in these microorganisms.

[Phage pf16 interrelationships with Pseudomonas putida bacteria. I. Unstable transductants and mutants of Pseudomonas putida PfG1 resistant to phage pf16].

The frequencies of transduction of the chromosomal genes by pf16 in Pseudomonas putida PgG1 are dependent on the marker transduced and unpredictable. Histidine and isoleucine-valine positive transductants, which are resistant to pf16, have been selected in the crosses with low (about 10(-8) for phage units) frequencies of transduction. Some of these transductants carry new mutations.

[Control of plasmid incompatibility: characteristics of the bireplicon hybrid pAS8 and its deletion mutants].

The phenomenon of incompatibility has been investigated using deletion mutants of hybrid bireplicon plasmid pAS8. The hybrid pAS8 displays incompatibility specific for both components of its structure. In contrast to P-specificity of pAS8, functions of ColE1-specificity are not effectively expressed.

Mapping of RP4 plasmid using deletion mutants of pAS8 hybrid (RP4--ColE1).

We used the hybrid plasmid pAS8 in order to conduct the genetic analysis of RP4 plasmid. The presence of two replicons in the hybrid plasmid permitted to expand the spectrum of deletion mutants of RP4 isolated, which are capable to autonomous replication. The shortening of the hybrid plasmid was achieved by P22 transduction, by induction of deletion mutants using mitomycin C, as well as by seletion of Tra- mutants on the basis of resistance of cells to P-specific phages.

[Genetic and heteroduplex analysis of deletion mutants of plasmid pAS8].

Heteroduplex analysis of deletion mutants of plasmid pAS8 permitted to construct a physical map and to elaborate in greater detail the genetic map of RP4 plasmid. The correlation between the ability of mutants to replicate in cells lacking functional DNA-polymerase I and the length of the deleted segment permitted to map rep genes of RP4 on DNA region with coordinates 9.8-17.

[Effect of amino acid substitutions in the polypeptide chain of DNA-polymerase on manifestation of the mutator effect].

Thin map of gene 43, controlling the synthesis of T4 DNA polymerase, is obtained by mapping experiments performed with 39 amber mutants, and is used for analysis of the sites of DNA polymerase gene from the point of view of displaying the mutator effect. The mutant sites studied possessed different reaction on amino acid substitutions in the polypeptide chain of the enzyme. Most of sites of the DNA polymerase gene, with the exception of two "supersensitive", responsed only on the apparent type of the amino acid substitutions: the mutator effect of amber mutations, which are located at these sites, was exhibited only in the case of insertion of the definite amino acid in the respective point of polypeptide chain.