Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Cosmids”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 649 records · Page 36Linked to original sources
PubMed1997

A 3-Mb contig from D11S987 to MLK3, a gene-rich region in 11q13.

We have combined genetic, radiation-reduced somatic cell hybrid (RRH), fluorescent in situ hybridization (FISH), and physical mapping methods to generate a contig of overlapping YAC, PAC, and cosmid clones corresponding to > 3 continuous Mb in 11q13. A total of 15 STSs [7 genes (GSTP1, ACTN, PC, MLK3, FRA1, SEA, HNP36), 4 polymorphic loci (D11S807, D11S987, GSTP1, D11S913), 3 ESTs (D11S1956E, D11S951E, and W1-12191), and 1 anonymous STS (D11S703)], mapping to three independent RRH segregation groups, identified 26 YAC, 7 PAC, and 16 cosmid clones from the CGM, Roswell Park, CEPH Mark I, and CEPH MegaYAC YAC libraries, a 5 genome equivalent PAC library, and a chromosome II-specific cosmid library. Thirty-six Alu-PCR products derived from 10 anonymous bacteriophage lambda clones, a cosmid containing the polymorphic marker D11S460, or STS-positive YAC or cosmid clones were identified and used to screen selected libraries by hybridization, resulting in the identification of 19 additional clones. The integrity and relative position of a subset of clones was confirmed by FISH and were found to be consistent with the physical and RRH mapping results. The combination of STS and Alu-PCR-based approaches has proven to be successful in attaining contiguous cloned coverage in this very GC-rich region, thereby establishing for the first time the absolute order and distance between the markers: CEN-MLK3-(D11S1956E/D11S951E/W1-12191)-FRA1-D 11S460-SEA-HNP36/ D11S913-ACTN-PC-D11S703-GSTP1-D11S987-TEL.

Base Sequence↗
PubMed1997

Intracellular carbonic anhydrase is essential to photosynthesis in Chlamydomonas reinhardtii at atmospheric levels of CO2. Demonstration via genomic complementation of the high-CO2-requiring mutant ca-1.

Genomic complementation of the high-CO2-requiring mutant ca-1-12-1C of Chlamydomonas reinhardtii was achieved by transformation with DNA pools from an indexed cosmid library of wild-type genomic DNA. Transformation of mutant cells with cosmid DNA from two microtiter plates in the library produced colonies that grew phototrophically at atmospheric CO2 levels. Transformations with cosmid DNA from each of the rows and files of the two plates pinpointed one well in each plate with a cosmid bearing the targeted gene. Sequencing of cosmid subclones revealed a gene encoding a recently identified C. reinhardtii chloroplast carbonic anhydrase (CAH3). Transformations with chimeric constructs combining different portions of the wild-type and mutant genes indicated the presence of a mutation in the 5'-half of the gene. Comparison of mutant and wild-type gene sequences in this region revealed a G-to-A substitution in the mutant gene, which produced a nonsense codon. The data presented demonstrate that the carbonic anhydrase produced from the CAH3 gene is essential to the inorganic carbon-concentrating mechanism in C. reinhardtii and that genomic complementation can be a facile and efficient means for isolating genes associated with defects affecting photosynthesis and other physiological processes in this eukaryotic green alga.

Amino Acid Sequence↗
PubMed1995

Cloning and characterization of the genes for p-nitrobenzoate degradation from Pseudomonas pickettii YH105.

Pseudomonas pickettii YH105 was isolated for its ability to utilize p-nitrobenzoate as the sole source of carbon, nitrogen, and energy. Degradation of p-nitrobenzoate by this strain proceeds through a reductive route as evidenced by the accumulation of ammonia in the culture medium during growth on p-nitrobenzoate. Enzyme assays and high-performance liquid chromatography (HPLC) analysis of culture supernatants indicate that p-nitrobenzoate is degraded through p-hydroxylaminobenzoate and protocatechuate. In order to clone the genes responsible for the initial steps in the catabolic pathway, a cosmid library was constructed with P. pickettii YH105 genomic DNA. The library was screened for clones capable of transforming p-nitrobenzoate to protocatechuate, using a plate assay specific for diphenolic compounds. HPLC analysis of culture supernatants confirmed that the cosmid clones did indeed produce protocatechuate from p-nitrobenzoate. Five positive cosmid clones that possessed this activity were identified. Restriction digests of the cosmid clones indicated that all of the clones had two EcoRI fragments in common (3.9 and 1.0 kb). One of these cosmid clones, designated pGJZ1601, was chosen for further analysis. Subcloning and activity assay experiments localized the genes responsible for the conversion of p-nitrobenzoate to protocatechuate to a 1.4-kb SalI-SphI DNA fragment. Further subcloning experiments localized the gene coding for p-nitrobenzoate reductase, responsible for the first enzymatic step in the catabolic pathway, to a 0.8-kb SalI-ApaI DNA fragment. The gene for the second step in the catabolic pathway, coding for hydroxylaminolyase, was located adjacent to the gene for the p-nitrobenzoate reductase.

Biodegradation, Environmental↗
PubMed1995

High-resolution alignment of a 1-megabase-long genome region of three strains of Rhodobacter capsulatus.

A detailed restriction map of the genome of Rhodobacter capsulatus SB1003 was constructed recently by using an ordered set of overlapping cosmids. Pulsed-field gel electrophoresis-generated restriction patterns of the chromosomes of 14 other R. capsulatus strains were compared. Two of them, St. Louis and 2.3.1, were chosen for high-resolution alignment of their genomes with that of SB1003. A 1-Mb segment of the R. capsulatus SB1003 cosmid set was used as a source of ordered probes to group cosmids from the other strains. Selected cosmids were linked into one 800-kb contig and two smaller contigs of 100 kb each. EcoRV and BamHI restriction maps of the newly ordered cosmids were constructed by using lambda terminase. Long-range gene order in the new strains was mainly conserved for the regions studied. However, one large genome rearrangement inverted a 470-kb DNA fragment of the St. Louis strain between the rrnA and rrnB operons. A 50-kb deletion covering three SB1003 probes was found in strain 2.3.1 near rrnB. Conservation of about 50% of the positions of restriction sites in all these strains and nearly 80% for the pair 2.3.1- St. Louis made it possible to produce high-resolution alignment of the contiguous 800-kb genome segment. Ten deletions of 2 to 27 kb, one 30-kb inversion, and three translocations were found in this region. Strong clustering of the positions of polymorphic restriction sites was observed. For a 50-kb size interval, two patterns of the distribution of restriction sites were found, one with about 90% and the other with 5 to 30% conservation of sites. This structure may be explained by independent acquisition of these divergent regions from other Rhodobacter strains.

Chromosome Inversion↗
PubMed1996

Allelic exchange in Mycobacterium tuberculosis with long linear recombination substrates.

Genetic studies of Mycobacterium tuberculosis have been greatly hampered by the inability to introduce specific chromosomal mutations. Whereas the ability to perform allelic exchanges has provided a useful method of gene disruption in other organisms, in the clinically important species of mycobacteria, such as M. tuberculosis and Mycobacterium bovis, similar approaches have thus far been unsuccessful. In this communication, we report the development of a shuttle mutagenesis strategy that involves the use of long linear recombination substrates to reproducibly obtain recombinants by allelic exchange in M. tuberculosis. Long linear recombination substrates, approximately 40 to 50 kb in length, were generated by constructing libraries in the excisable cosmid vector pYUB328. The cosmid vector could be readily excised from the recombinant cosmids by digestion with PacI, a restriction endonuclease for which there exist few, if any, sites in mycobacterial genomes. A cosmid containing the mycobacterial leuD gene was isolated, and a selectable marker conferring resistance to kanamycin was inserted into the leuD gene in the recombinant cosmid by interplasmid recombination in Escherichia coli. A long linear recombination substrate containing the insertionally mutated leuD gene was generated by PacI digestion. Electroporation of this recombination substrate containing the insertionally mutated leuD allele resulted in the generation of leucine auxotrophic mutants by homologous recombination in 6% of the kanamycin-resistant transformants for both the Erdman and H37Rv strains of M. tuberculosis. The ability to perform allelic exchanges provides an important approach for investigating the biology of this pathogen as well as developing new live-cell M. tuberculosis-based vaccines.

Alleles↗
PubMed2002

Rapid construction of adenoviral vectors by lambda phage genetics.

Continued improvements of adenoviral vectors require the investigation of novel genome configurations. Since adenovirus can be generated directly by transfecting packaging cell lines with viral genomes isolated from plasmid DNA, it is possible to separate genome construction from virus production. In this way failure to generate a virus is not associated with an inability to generate the desired genome. We have developed a novel lambda-based system that allows rapid modification of the viral genome by double homologous recombination in Escherichia coli. The recombination reaction and newly generated genome may reside in a recombination-deficient bacterial host for enhanced plasmid stability. Furthermore, the process is independent of any restriction endonucleases. The strategy relies on four main steps: (i) homologous recombination between an adenovirus cosmid and a donor plasmid (the donor plasmid carries the desired modification[s] and flanking regions of homology to direct its recombination into the viral genome); (ii) in vivo packaging of the recombinant adenoviral cosmids during a productive lambda infection; (iii) transducing a recombination-deficient E. coli lambda lysogen with the generated lysate (the lysogen inhibits the helper phage used to package the recombinant andenoviral cosmid from productively infecting and destroying the host bacteria); (iv) effectively selecting for the desired double-recombinant cosmid. Approximately 10,000 double-recombinant cosmids are recovered per reaction with essentially all of them being the correct double-recombinant molecule. This system was used to generate quickly and efficiently adenoviral genomes deficient in the E1/E3 and E1/E3/E4 regions. The basis of this technology allows any region of the viral genome to be readily modified for investigation of novel configurations.

Adenoviridae↗
PubMed1988

Molecular cloning and biological characterization of a human gene, ERCC2, that corrects the nucleotide excision repair defect in CHO UV5 cells.

The UV-sensitive Chinese hamster ovary (CHO) cell line UV5, which is defective in the incision step of nucleotide excision repair, was used to identify and clone a complementing human gene, ERCC2, and to study the repair process. Genomic DNA from a human-hamster hybrid cell line was sheared and cotransferred with pSV2gpt plasmid DNA into UV5 cells to obtain five primary transformants. Transfer of sheared DNA from one primary transformant resulted in a secondary transformant expressing both gpt and ERCC2. The human repair gene was identified with a probe for Alu-family repetitive sequences. For most primary, secondary, and cosmid transformants, survival after UV exposure showed a return to wild-type levels of resistance. The levels of UV-induced mutation at the aprt locus for secondary and cosmid transformants varied from 50 to 130% of the wild-type level. Measurements of the initial rate of UV-induced strand incision by alkaline elution indicated that, whereas the UV5 rate was 3% of the wild-type level, rates of cosmid-transformed lines were similar to that of the wild type, and the secondary transformant rate was about 165% of the wild-type rate. Analysis of overlapping cosmids determined that ERCC2 is between 15.5 and 20 kilobases and identified a closely linked gpt gene. Cosmids were obtained with functional copies of both ERCC2 and gpt. ERCC2 corrects only the first of the five CHO complementation groups of incision-defective mutants.

Animals↗
PubMed1989

Genome mapping in halobacteria.

The goal of our research is to produce an ordered set of cosmid clones for each of several species of halobacteria for use in physical and genetic mapping. These maps will answer questions about genome evolution and about gene organization and regulation in this archaebacterial lineage. Progress in cloning and mapping the genome of Halobacterium volcanii DS2 (synonym Haloferax volcanii DS2) is reported. Overlapping cosmids are recognized by a strategy which makes use of the distinctive restriction fragments around relatively rare restriction sites. Each site recognized by the infrequently cutting restriction enzymes is a landmark from which to identify different regions of the genome. The main advantage of this strategy is that only a small overlap (10-20%) between cosmid clones is required, resulting in a correspondingly small number of cosmid clones to be analyzed. The certainty of overlap is high, and computation is simple. The final 5-10% of each genome is cloned, linked, and identified by chromosome walking methods. Hybridization of cloned homologous or heterologous genes and of stable RNAs to the minimal cosmid set localizes these genes on the physical map. Additional genes have been and will be cloned by complementation of auxotrophic mutants, or as determinants of resistance to antibiotics.

Chromosome Mapping↗
PubMed1997

Segmental homology among cattle (Bos taurus), Indian muntjac (Muntiacus muntjak vaginalis), and Chinese muntjac (M. reevesi) karyotypes.

In an attempt to examine homologies between Indian and Chinese muntjac karyotypes at a subchromosomal level, five bovine cosmids were comparatively mapped by heterologous fluorescence in situ hybridization (FISH). In the Indian muntjac (2n = 6) all cosmids mapped to chromosome 1, whereas in the Chinese muntjac (2n = 46) two cosmids mapped to chromosome 3 and one cosmid each mapped to chromosomes 1, 7, and 17. These markers have maintained their intrachromosomal position relative to a centromere/telomere axis in cattle and in Chinese and Indian muntjac chromosomal arms. Our results corroborate the tandem-fusion hypothesis for muntjac karyotypic evolution and establish orientational homology between the involved Chinese muntjac chromosomes and the discrete segments on Indian muntjac chromosome 1. Furthermore, our data disclose regional homologies between cattle and muntjac genomes and demonstrate the validity of intergeneric cosmid-FISH for investigations on karyotype evolution.

Animals↗
PubMed1993

[Application of the chromosome sorting technique to the human genome analysis].

The flow-sorted human chromosome is an ideal DNA source for genome analysis research, particularly gene mapping and construction of cosmid library. For gene mapping, 50,000 chromosomes each were sorted on a membrane filter and hybridized with a radio-labeled gene probe. Using this Spot Blot Hybridization method in combination with FISH, we have mapped more than 30 new genes including 6 neuron-specific ones. For construction of a chromosome-specific cosmid library, nearly 10 million chromosomes were flow-sorted. The chromosome 21-specific cosmid library consisting of 33,000 clones and the 22pter-q11 region-specific library consisting of 9,200 clones were established. Grouping of cosmid clones from the library using high density replica filters and YAC DNAs to make cosmid contigs within specific regions is under way.

Base Sequence↗
PubMed1995

Construction of a complete genomic library of Saccharomyces cerevisiae and physical mapping of chromosome XI at 3.7 kb resolution.

A consortium of European laboratories has been organized to systematically sequence the genome of Saccharomyces cerevisiae. As part of the BIOTECH program aimed at sequencing chromosomes XI and II, we have constructed a total genomic library of yeast strain FY1679 (a direct S288C derivative) into cosmid vectors pWE15 and pOU61cos. Primary clones from four independent libraries totalling 190 genome equivalents have been stored at -80 degrees C. A subset of 1939 independent clones (six genome equivalents) was hybridized using purified chromosomes XI and X as probes. A total of 147 chromosome XI-specific cosmid clones was used to construct the physical map of that chromosome. Mapping methods included a combination of classical bottom-up strategies (fingerprinting, hybridizations) and a novel top-down strategy using I-SceI chromosome fragmentation. The 147 cosmid clones form a unique contig covering the entire chromosome XI (666 kb) with the sole exceptions of the (C1-3A)n repeats of the telomeres. Colinearity of cosmid inserts with yeast DNA was directly verified. A complete EcoRI map of chromosome XI was deduced from partial overlaps of cosmids and used for the sequencing program. Comparison of this map with the genetic map shows unexpected divergences that have been solved by subsequent genetic analysis, yet underline the necessity of independent physical mapping in genome projects.

Base Sequence↗
PubMed1993

Evidence for a gene cluster involving trichothecene-pathway biosynthetic genes in Fusarium sporotrichioides.

Two overlapping cosmid clones (Cos1-1 and Cos9-1) carrying the Tox5 gene were isolated from a library of F. sporotrichioides strain NRRL 3299 genomic DNA. These cosmids were used to transform three T-2 toxin-deficient mutants that are blocked at different steps in the trichothecene pathway. Both cosmids restored T-2 toxin production to Tox3-1- or Tox4-1- mutants but neither restored T-2 toxin production to a Tox1-2- mutant. The production of T-2 toxin by the complemented Tox3-1- and Tox4-1- mutants, as well as the production of diacetoxycirpenol by the cosmid-transformed Tox1-2- mutant, were 2- to 10-fold higher than in strain NRRL 3299. In addition, those transformants carrying Cos9-1 produced significantly higher levels of trichothecenes than transformants carrying Cos1-1. Two different DNA fragments (FSC13-9 and FSC14-5), representing the region of overlap between the cosmid clones, were isolated. These fragments specifically complemented either the Tox3-1- mutant (FSC14-5) or the Tox4-1- mutant (FSC13-9). The trichothecene-production phenotype of these transformants was similar to NRRL 3299. These results suggest that two or more genes involved in the biosynthesis of trichothecenes are closely linked to Tox5.

Cloning, Molecular↗
PubMed1995

Cloning of genes required for hypersensitivity and pathogenicity in Pseudomonas syringae pv. aptata.

A genomic library of Pseudomonas syringae pv. aptata strain NCPPB 2664, which causes bacterial blight of sugar beet, lettuce and other plants, was constructed in the cosmid vector pCPP31. The 13.4 kb EcoRI fragment of the cosmid pHIR11, containing the hrp (hypersensitive response and pathogenicity) gene cluster of the closely related bacterium Pseudomonas syringae pv. syringae strain 61, was used as a probe to identify a homologous hrp gene cluster in P. syringae pv. aptata. Thirty of 2500 cosmid clones, screened by colony hybridization, gave a strong hybridization signal with the probe, but none of these conferred to the non-pathogenic bacterium, Pseudomonas fluorescens, the ability to elicit the hypersensitive response (HR) in tobacco. Southern blot analysis of EcoRI-digested genomic DNA of P. syringae pv. aptata showed hybridizing bands of 12 kb and 4.4 kb. Only a 12 kb fragment hybridized in digests of the cosmids. Cosmid clone pCPP1069 was mutagenized with Tn10-minitet and marker-exchanged into the genome of P. syringae pv. aptata. Three resulting prototrophic mutant strains failed to elicit the HR in tobacco and to cause disease in lettuce. The DNA flanking the Tn10-minitet insertions from mutated derivatives of pCPP1069 hybridized with the 10.6 kb Bg/II fragment of pHIR11. These results indicate that P. syringae pv. aptata harbours hrp genes that are similar to, but arranged differently from, homologous hrp genes of P. syringae pv. syringae.

Blotting, Southern↗
PubMed1985

Cloning of the DNA repair genes mtcA, mtcB, uvsC, uvsD, uvsE and the leuB gene from Deinococcus radiodurans.

A gene library from Deinococcus radiodurans has been constructed in the cosmid pJBFH. A 51.5-kb hybrid cosmid, pUE40, that transduced Escherichia coli HB101 from leucine dependence to independence was selected, and a 6.9-kb fragment which carried the leuB gene from D. radiodurans was subcloned into the EcoRI site of pAT153. The DNA repair genes mtcA, mtcB, uvsC, uvsD and uvsE, which code for two D. radiodurans UV endonucleases were identified by transforming appropriate repair-deficient mutants of D. radiodurans to repair proficiency with DNA derived from the gene library. Hybrid cosmid pUE50 (37.9 kb) containing an insert carrying both the mtcA and mtcB genes was selected and 5.6- and 2.7-kb DNA fragments carrying mtcA and mtcB, respectively, i.e., the genes that code for UV endonuclease alpha, were subcloned into the EcoRI site of pAT153. The three genes uvsC, uvsD and uvsE, that code for UV endonuclease beta, were all present in the 46.0-kb hybrid cosmid pUE60. The uvsE gene in a 12.2-kb fragment was subcloned into the HindIII site of pAT153 and the size of the insert reduced to 6.1 kb by deletion of a 6.7-kb fragment from the hybrid plasmid pUE62. None of the uvs genes introduced into the UV-sensitive E. coli CSR603 (uvrA-) was able to complement its repair defect. The mtcA, uvsC, uvsD and uvsE genes were found in the 52.5-kb hybrid cosmid pUE70. It is concluded that the DNA repair genes mtcA, mtcB, uvsC, uvsD and uvsE are located within an 83.0-kb fragment of the D. radiodurans genome.

Chromosome Mapping↗
PubMed1995

Structure, sequence and location of the UQCRFS1 gene for the human Rieske Fe-S protein.

We have identified and studied the chromosomal location of the human Rieske Fe-S protein-encoding gene UQCRFS1. Mapping by hybridization to a panel of monochromosomal hybrid cell lines indicated that a UQCRFS1 partial cDNA was derived from either chromosome 19 or 22. By screening a human chromosome 19 specific genomic cosmid library with a probe from this cDNA sequence, we identified a corresponding cosmid. Portions of this cosmid were sequenced directly. The exon, exon:intron junction and flanking sequences verified that this cosmid contains the genomic locus. Fluorescent in situ hybridization (FISH) was performed to localize this cosmid to chromosome band 19q12.

Amino Acid Sequence↗
PubMed2001

Arabidopsis UVH3 gene is a homolog of the Saccharomyces cerevisiae RAD2 and human XPG DNA repair genes.

To identify mechanisms of DNA repair in Arabidopsis thaliana, we have analyzed a mutant (uvh3) which exhibits increased sensitivity to ultraviolet (UV) light, H2O2 and ionizing radiation and displays a premature senescence phenotype. The uvh3 locus was mapped within chromosome III to the GL1 locus. A cosmid contig of the GL1 region was constructed, and individual cosmids were used to transform uvh3 mutant plants. Cosmid N9 was found to confer UV-resistance, H2O2-resistance and a normal senescence phenotype following transformation, indicating that the UVH3 gene is located on this cosmid and that all three phenotypes are due to the same mutation. Analysis of cosmid N9 sequences identified a gene showing strong similarity to two homologous repair genes, RAD2 (Saccharomyces cerevisiae) and XPG (human), which encode an endonuclease required for nucleotide excision repair of UV-damage. The uvh3 mutant was shown to carry a nonsense mutation in the coding region of the AtRAD2/XPG gene, thus revealing that the UVH3 gene encodes the AtRAD2/XPG gene product. In humans, the homologous XPG protein is also involved in removal of oxygen-damaged nucleotides by base excision repair. We discuss the possibility that the increased sensitivity of the uvh3 mutant to H2O2 and the premature senescence phenotype might result from failure to repair oxygen damage in plant tissues. Finally, we show that the AtRAD2/XPG gene is expressed at moderate levels in all plant tissues.

Amino Acid Sequence↗
PubMed1983

Construction of a Rhizobium japonicum gene bank and use in isolation of a hydrogen uptake gene.

A gene bank of Rhizobium japonicum DNA was constructed by using the broad host range conjugative cosmid pLAFR1. Eighty-three percent of the clones in the bank contained cosmids with insert DNA averaging 22.6 kilobase pairs in length. A series of cosmids containing a hydrogen uptake (hup) gene was identified by transferring the gene bank into a H(2) uptake-negative (Hup(-)) R. japonicum point mutant (PJ17nal) and screening tetracycline-resistant colonies for the ability to grow chemolithotrophically and to reduce methylene blue in a recently devised colony assay. Hup(+) transconjugants arose at a frequency of approximately 6 x 10(-3). Plasmid DNAs from II of the Hup(+) transconjugants were isolated and used to transform Escherichia coli. EcoRI digests of all plasmids isolated from Hup(+) transconjugants had three DNA fragments in common. Eight of the E. coli transformants containing hup gene cosmids were conjugated with PJ17nal and another Hup(-) point mutant, PJ18nal. All PJ17nal transconjugants were Hup(+). The frequency of Hup(+) transconjugants with PJ18nal was approximately 10(-3). The results indicate that the hup gene cosmids may contain one gene and a portion of another.

Journal Article↗
PubMed1988

Isolation of genes involved in nodulation competitiveness from Rhizobium leguminosarum bv. trifolii T24.

Rhizobium leguminosarum bv. trifolii T24 produces a potent anti-rhizobial compound, trifolitoxin, and exclusively nodulates clover roots when in mixed inoculum with trifolitoxin-sensitive strains of R. leguminosarum bv. trifolii [Schwinghamer, E. A. & Belkengren R. P. (1968) Arch. Mikrobiol. 64, 130-145]. In the present study, the isolation of trifolitoxin production and resistance genes is described. A cosmid genomic library of T24 was prepared in pLAFR3. No trifolitoxin expression was observed in the resulting Escherichia coli cosmid clones. One cosmid clone was identified that restored trifolitoxin production and nodulation competitiveness in three nonproducing mutants of T24. The recombinant plasmid from this cosmid clone, pTFX1, also conferred trifolitoxin production and resistance when transferred to symbiotically effective strains of R. leguminosarum bvs. trifolii, phaseoli, and viceae. Cosmid pTFX1 also conferred expression of trifolitoxin production when present in strains of Rhizobium meliloti and Agrobacterium tumefaciens. No trifolitoxin expression was observed in strains of Bradyrhizobium japonicum or Rhizobium sp. (cowpea) with pTFX1. Southern blot analysis with a biotinylated pTFX1 probe did not suggest that these genes were plasmid-borne. Transfer of pTFX1 to T24 or its derivatives resulted in 6- to 10-fold higher level of trifolitoxin production than wild-type T24.

Journal Article↗