Transposon Tn951 (TnLac) is defective and related to Tn3

Summary Tn951 is flanked by two perfect inverted repeats of 41 bp which include the 38 bp sequence of the IR of Tn3. Tn951 also contains the last 100 bp of the tnpA gene but with at least two mutations. However, beyond nucleotide 137 the sequences diverge and hybridization experiments show that Tn951 lacks at least the first two thirds of the tnpA gene.In agreement with these observations Tn951 does not transpose by itself at a detectable frequency but can be complemented by the tnpA gene of Tn801 or Tn3. Tn501, Tn1721 and gamma delta do not complement Tn951 transposition.Transposition of Tn951 duplicates 5 bp of target DNA sequence.     

Abundance of Tn3, Tn21, and Tn501 transposase (tnpA) sequences in bacterial community DNA from marine environments.

The occurrence of the tnpA genes of the transposons Tn3, Tn21, and Tn501 was assessed in total bacterial community DNA isolated from different marine environments. The PCR technique was employed, together with most probable number statistics, to determine the abundance of the target tnpA genes. All three genes could be detected, and the Tn21 tnpA sequences predominated in all samples. The smallest amount of total community DNA in which the Tn21 tnpA sequence could be detected was 0.037 ng, and on the basis of our results, we estimated that this sequence was present in 1 of 1,000 to 10,000 bacteria. Hybridization of the PCR products with the respective tnpA probes verified the Tn21 and Tn501 tnpA sequences but only some of the Tn3 tnpA amplification products. The distribution and dissemination of transposons in natural bacterial communities are discussed.     

Streptococcus faecalis R plasmid pJH1 contains an erythromycin resistance transposon (Tn3871) similar to transposon Tn917

The R plasmid pJH1 contains a 5.1-kilobase transposon ( Tn3871 ) that mediates inducible resistance to erythromycin. Three AvaI digestion fragments from this transposon are identical in size to and homologous with three AvaI-derived fragments from the previously described erythromycin resistance transposon Tn917 . These three DNA fragments account for greater than 90% of both transposons.     

Characterization of the transposons Tn1822 (Tc) and Tn1824 (TpSm) and the light they throw on the natural spread of resistance genes

The transposons Tn1822 (Tc) and Tn1824 (TpSm) described in this paper are very similar or identical to the transposons Tn1771 and Tn1721 (Tc), and Tn7 (TpSm), respectively. They were isolated from different sources and replicons, however, suggesting a wide distribution of these types of transposons. The method used to isolate and characterize the transposons, involving a temperature-sensitive P1 phage as their transient vehicle, is particularly suitable for epidemiological studies on transposon distribution.     

Characterization of Tn916S, a Tn916-like element containing the tetracycline resistance determinant tet(S)

We have characterized a transferable tetracycline resistance (Tcr) element from a Streptococcus intermedius isolate. The gene responsible for this resistance was identified by PCR and Southern hybridization as tet(S). Furthermore, the genetic support for this determinant was shown to be a conjugative transposon closely related to Tn916. This element has been designated Tn916S.     

Expression of a Lactose Transposon (Tn951) in Zymomonas mobilis

The potential utility of Zymomonas mobilis as an organism for the commercial production of ethanol would be greatly enhanced by the addition of foreign genes which expand its range of fermentable substrates. We tested various plasmids and mobilizing factors for their ability to act as vectors and introduce foreign genes into Z. mobilis CP4. Plasmid pGC91.14, a derivative of RP1, was found to be transferred from Escherichia coli to Z. mobilis at a higher frequency than previously reported for any other plasmids. Both tetracycline resistance and the lactose operon from this plasmid were expressed in Z. mobilis CP4. Plasmid pGC91.14 was stably maintained in Z. mobilis at 30°C but rapidly lost at 37°C.     

Integration of transposon Tn10 into phage L (Salmonella typhimurium)

Transposon Tn10 was transposed into phage L (Salmonella typhimurium) from F'ts114lac+zzf::Tn10 plasmid of strain TT629 (Chumely et al. 1979). Phage L with the insertion Tn10 (L::Tn10-8) was isolated in the form of a prophage in the lysogenic strain S. typhimurium LT2-18 (L::Tn10-8), in which it can be induced with UV light. The phage induced in this way is defective; however, it forms plaques at a multiplicity of infection (moi) greater than one and transduces the tetracycline-resistance determinant to tetracycline-sensitive cells. Analysis of its DNA by restriction endodeoxyribonucleases revealed insertion of the intact transposon Tn10 of 9300 bp in the E fragment, formed during the action of EcoRI, at a distance of 16,800 bp from the pac site.     

Tn5253, the pneumococcal omega (cat tet) BM6001 element, is a composite structure of two conjugative transposons, Tn5251 and Tn5252.

Tn5253, carrying tetracycline and chloramphenicol resistance determinants, is a 65.5-kb conjugative transposon originally detected in the chromosome of Streptococcus pneumoniae BM6001. We have identified an 18-kb segment of DNA carrying the tet determinant within Tn5253 to be an independent conjugative transposon when removed from the context of the larger element. In vivo deletion of this DNA segment, now termed Tn5251, from within Tn5253 did not affect the conjugative transposition properties of the remaining sequences. Thus, Tn5253 is a composite element of two conjugative structures: Tn5252, constituting the sequences beyond Tn5251 within Tn5253, and Tn5251. The transfer properties of Tn5252 and Tn5251 suggest that these may belong to two different classes of mobile elements even though they were initially found associated. The notion that a tet-carrying transposon like Tn5251 may have been the ancestral element in the evolution of the larger streptococcal conjugative transposons must be reevaluated in the light of present observations.     

Transposition of Tn951 (Tnlac) and cointegrate formation are thermosensitive processes

The frequency of transposition of Tnlac to pGC200, an IncFII R plasmid, increased during the storage of the host strain. This result is explained by the fact that the transpositional event is temperature-dependent: it occurred readily when the host strain was grown at 30 degrees C but it was nearly undetectable when the host strain was grown and kept at 37 degrees C. Fusions between two different plasmids carrying Tnlac with pGC200 were also thermosensitive, suggesting a relation between cointegrate formation and transposition. Lactose did not influence the frequency of transposition of Tnlac.     

The R46 site-specific recombination system is a homologue of the Tn3 and gamma delta (Tn1000) cointegrate resolution system

The nucleotide sequence of the R46 site-specific recombination system has been determined. The organization of the recombination gene (perR46) and the site at which it acts (per site), together with the extensive sequence homology displayed with the tnpR genes and res sites of the transposons Tn3 and gamma delta (Tn1000), suggests that they have been derived from a Tn3-like element. These site-specific recombination functions of R46 play a role in plasmid maintenance.     

Intramolecular transposition by a synthetic IS50 (Tn5) derivative.

We report the formation of deletions and inversions by intramolecular transposition of Tn5-derived mobile elements. The synthetic transposons used contained the IS50 O and I end segments and the transposase gene, a contraselectable gene encoding sucrose sensitivity (sacB), antibiotic resistance genes, and a plasmid replication origin. Both deletions and inversions were associated with loss of a 300-bp segment that is designated the vector because it is outside of the transposon. Deletions were severalfold more frequent than inversions, perhaps reflecting constraints on DNA twisting or abortive transposition. Restriction and DNA sequence analyses showed that both types of rearrangements extended from one transposon end to many different sites in target DNA. In the case of inversions, transposition generated 9-bp direct repeats of target sequences.     

Tn5-induced mutations affecting sulfur-oxidizing ability (Sox) of Thiosphaera pantotropha

Mutants of Thiosphaera pantotropha defective in chemolithoautotrophic growth were obtained by transpositional mutagenesis with Tn5 coding for kanamycin resistance. The suicide vehicle for introducing Tn5 to T. pantotropha was pSUP5011 harbored by Escherichia coli. Kanamycin-resistant isolates were screened for the inability to grow with reduced sulfur compounds (Sox-). Four classes of Sox- mutants were obtained. Three were of different pleiotropic phenotypes: (i) unable to grow with formate, nitrate, and xanthine; (this class strongly suggested the involvement of a molybdenum cofactor in inorganic sulfur-oxidizing ability); (ii) no growth with hydrogen; (iii) slight growth with hydrogen and formate. Two plasmids, pHG41 (about 450 kilobase pairs) and pHG42 (110 kilobases), were identified in lysates of T. pantotropha. In one Sox- mutant pHG41 could not be detected. Revertant analysis suggested that pHG41 and pHG42 were not involved in the Sox character.     

Plasmid Reference Center Registry of transposon(Tn) and insertion sequence (IS) allocations through December 1986

The current entries of transposable genetic elements (both Tn and IS) filed with the Plasmid Reference Center are tabulated. These include Tn entries 3601-4550 [entries 1-3600 are listed in Lederberg, Gene 16 (1981) 59-61 and 18 (1982) 366] and all the filed IS entries.     

Inversions and deletions generated by a mini-gamma delta (Tn1000) transposon.

Intramolecular transposition by an engineered derivative of the transposon gamma delta (Tn1000) is described. This 1-kb element contains inverted repeats of the 40 bp of the delta end of gamma delta, bracketing a kan gene, but it contains no resolution site. Transposition was analyzed in two plasmids; one contained two contraselectable (conditional lethal) genes (thyA and sacB) adjacent to the mini-gamma delta element in a 13.0-kb pBR322/pUC-based two-component plasmid (a heterodimer), and the other contained a different contraselectable gene (strA [rpsL]) in a 13.2-kb three-component plasmid (a heterotrimer). Selection for loss of function of a single contraselectable gene yielded inversions and deletions. Each inversion plasmid was 1 kb larger than the parent plasmid: it had a second copy of mini-gamma delta inserted in the contraselected gene, with that copy plus the intervening segment inverted, and the 5-bp target site duplicated. Each deletion plasmid was smaller than the parent plasmid and had a deletion that extended from one transposon end into or through the contraselected gene for distances of up to 9.4 kb. The frequencies of deletions versus inversions ending in a single target gene were similar, although overall, deletions outnumbered inversions because deletions, but not inversions, into sites beyond the contraselected gene inactivate it. This work also demonstrates that thyA (which encodes thymidylate synthetase) is a useful contraselectable marker.     

Complementation of transposition of tnpA mutants of Tn3, Tn21, Tn501, and Tn1721

The transposons Tn21, Tn501, and Tn1721 are related to Tn3. Transposition-deficient mutants (tnpA) of these elements were used to test for complementation of transpostion. Transposition of tnpA mutants of Tn501 and Tn1721 was restored by the presence in trans of Tn21, Tn501, and Tn1721, but transposition of a tnpA mutant of Tn21 was restored in trans only by Tn21 itself. Tn3 did not complement transposition of Tn21, Tn501, or Tn1721, and these elements did not complement transposition of Tn3.