Germ line specific DNA and chromosomes of the ciliate Stylonychia lemnae

The variation in DNA content of the micronucleus (germinal nucleus) of Stylonychia lemnae and its relation to the number of chromosomes was examined. Different populations possess similar amounts of micronuclear DNA but there are differences of ±30% between clones of the same population. However, the DNA content varies by about 100% in the micronuclei during the lifetime of a clone. The haploid micronucleus contains 35 or 36 chromosomes which persist in the developing macronucleus anlagen and grow to giant chromosomes. Besides this remaining subset, the micronucleus contains a variable number of germ line restricted chromosomes (mean about 140; range between 100 and 180). The somatic macronucleus eliminates these elements early in its development. The varying number of the germ line restricted chromosomes is responsible for the variation in the micronuclear DNA content.     

Growth of Nitrobacter by dissimilatoric nitrate reduction

Abstract Eight strains of the genus Nitrobacter grew under anaerobic conditions in the presence of nitrate. The growth was inhibited by nitrate concentrations above 0.5 mM. By a special culture technique inhibition caused by nitrite was abolished. Nitrate oxidizing cells grew in gas tight culture flasks as a biofilm on a gas-permeable silicone tubing. The biofilm allowed nitrate-reducing cells to grow at a low nitrite concentration. These cells grew either actively motile in the anaerobic medium, or in anaerobic zones of the biofilm. They produced nitrite and ammonia. Nitrogen balance calculations established a loss of inorganic nitrogen for 5 of 8 strains. This implies that nitrate-reducing cells produced furthermore volatile nitrogen compounds. N₂O was detected by gas chromatography.     

A mutant rat major histocompatibility haplotype showing a large deletion of class I sequences

The LEW.1LM1 inbred rat strain, which has been derived from a (LEW×LEW.1W) F₂ hybrid, carries a major histocompatibility (RT1) haplotype which is distinct from that of the LEW strain (RT1 ¹) in that certainRT1.C region-determined class I antigens are not expressed. Here we show that this phenotypic defect is due to genomic deletion of about 100 kb of theRT1.C region. Certain deleted DNA fragments have been cloned from the wild-type DNA into the EMBL4 vector. Five clones have been characterized and are shown to possess different restriction maps and to each carry a single stretch of class I cross-hybridizing sequences. Probes derived from the non-class I coding part of two clones detect fragments which are present in the wild-type but absent from thelm1 mutant. The type of deletion described here in the rat is discussed in the context ofH-2D/Q deletions in the mouse.     

Electrotransformation of intact and osmotically sensitive cells of Corynebacterium glutamicum

Summary Intact and osmotically sensitive cells of Corynebacterium glutamicum can be efficiently transformed by electroporation. This was shown by using the plasmid vector pUL-330 (5.2 kb), containing the kanamycin resistance gene of transposon Tn5. The following electric parameters yielded efficient transformation. For intact cells: one exponentially decaying field pulse with time constants and with initial field intensities of E ₀=35–40 kV cm^-1; prepulse temperature 20°C. Cell regeneration (survival) was 100%–80%. Transformation efficiency can be increased by an additional freeze and thaw cycle of the cells, prior to electroporation. Lysozyme treated cells (osmotically sensitive) were transformed with three successive pulses of E ₀=25–30 kV cm^-1. Cell regeneration under these conditions was found to be 20–30%. The optimum yield of transformants/g plasmid-DNA was 3×10³ for intact cells, 2×10⁴ for intact cells which were frozen and thawed twice and 7×10⁴ for osmotically sensitive cells if the cell suspension was pulsed at a cell density of 1–3×10⁸/ml and at a DNA concentration of 0.2 g/ml up to 2 g/ml. The data obtained for osmotically sensitive cells suggest that the temperature increase accompanying the electric field pulse enhances colony formation and transformation efficiency if the initial prepulse temperature is 20°C, although regeneration of electroporated C. glutamicum cells starts to decrease at temperatures20°C.     

Modification of a catalytically important residue of indoleglycerol-phosphate synthase from Escherichia coli

The active-site residues of indoleglycerol-phosphate synthase from Escherichia coli were tentatively localized by comparing crystallographic data with the amino acid identities among the known indoleglycerol-phosphate synthase sequences. To test the validity of the resulting model of catalysis one of the residues in the presumptive active site, Lys 55, was changed to serine using oligonucleotide-directed mutagenesis. The specificity constant kcat/Km of the mutant is 3 x 10(4)-times lower than that of the wild-type enzyme, due to a 60-fold decrease in kcat and a 450-fold increase in Km. This finding shows that Lys 55 is important for both catalysis and substrate binding.     

A new facultatively nitrite oxidizing bacterium,Nitrobacter vulgaris sp. nov.

A total of 17 facultatively lithoautotrophic strains of Nitrobacter were investigated. They all were found to be related on the species level by DNA hybridizations. The G+C content of DNA ranged between 58.9 and 59.9 mol %. The isolates originated from divers environments. The cells were 0.5–0.8×1.2–2.0 m in size and motile by one polar to subpolar flagellum. Cell-division normally occurred by budding. Polar caps of intracytoplasmic membranes as well as carboxysomes were present. The cells tended to excrete extracellular polymers forming aggregates or biofilms. Heterotrophic growth was slower than mixotrophic but often faster than litoautotrophic growth. In the presence of nitrite and organic substances the organisms often showed diphasic growth. First nitrite and then the organic material was oxidized. In the absence of oxygen growth was possible by dissimilatory nitrate reduction. Nitrite, nitric and nitrous oxide as well as ammonia were formed. Depending on growth conditions the generation times varied from 12 to 140 h. The new Nitrobacter spec. may be one of the most abundant nitrite-oxidizing bacteria in soils, fresh waters and natural as well as artificial stones. For this organism the name Nitrobacter vulgaris is proposed.The type strain is filed with the culture collection of the Institut für Allgemeine Botanik, Universität Hamburg, FRG.     

Degradation of 3-chlorobiphenyl by in vivo constructed hybrid pseudomonads

Abstract 3-Chlorobiphenyl-degrading bacteria were obtained from the mating between Pseudomonas putida strain BN10 and Pseudomonas sp. strain B13. Strains such as BN210 resulted from the transfer of the genes coding the enzyme sequence for the degradation of chlorocatechols from B13 into BN10, whereas B13 derivatives such as B131 have acquired the biphenyl degradation sequence from BN10. During growth of the hybrid strains on 3-chlorobiphenyl 90% chloride was released. Activities of phenylcatechol 2,3-dioxygenase, benzoate dioxygenase, catechol 1,2-dioxygenase, chloromuconate cyloisomerase and 4-carboxymethyl-enebut-2-en-4-olide hydrolase were found in 3-chlorobiphenyl-grown cells. The hybrid strains were found to convert some congeners of the Aroclor 1221 mixture such as mono- and dichloro-substituted biphenyls.     

The control of root, vegetative shoot and flower morphogenesis in tobacco thin cell-layer explants (TCLs)

Thin cell-layer explants (TCLs) have been proposed as favorable tissues for the study of root, vegetative shoot and flower formation. We tested the effects of pH, light quality, light quantity, and IBA and kinetin concentrations on the morphogenesis of TCLs cultured individually on a liquid medium. Alterations of the amounts of exogenously supplied IBA and kinetin were sufficient to induce the formation of roots, vegetative shoots and flowers on TCLs cultured on otherwise identical media. The type and number of organs formed were sensitive to the intensity of light (55, 75, 100 and 120 muEinsteins m-2 sec-1) under which TCLs were grown. Evidence was obtained that the effects of light on TCL morphogenesis were associated with photochemical degradation of IBA in the medium. Evaluation of the organogenesis that occurred in TCLs cultured on a medium containing a range of IBA and kinetin concentrations showed that the number and type of organs formed, and overall growth, were dependent upon the initial concentrations of auxin and cytokinin. We have developed the TCL culture system into a sensitive and reproducible bioassay for the study of morphogenesis. The advantages of using the TCL morphogenesis bioassay for the identification and study of molecules (e.g. cell wall oligosaccharides) that may regulate morphogenesis are discussed.     

Bioconversion of 3-chlorobenzoate to 2-chloromuconate controlled by on line HPLC

Summary Strain RD330 a transposon mutant of Alcaligenes eutrophus JMP134 was considered to be dienelactone hydrolase defective (Don et al. 1985). During a bioconversion experiment with 3CB (3-chlorobenzoate) 2CMA (2-chloro-cis,cis-muconate) was accumulated by RD330 with an overall amount of 31%, but no dienelactone could be detected. Enzyme tests revealed that both enzymes 2CMA-cycloisomerase and dienelactone-hydrolase were induced at low levels in RD330 by 3CB and its metabolites.The control of 3CB addition during the bioconversion experiment was performed by on line HPLC (high pressure liquid chromatography).