E.coli tRNALeu的提纯

本文报道一种E.coli tRNA^Leu简便而稳定的纯化方法。粗tRNA经过BD-Cellulose柱层析和聚丙烯酰胺凝胶电泳两个步骤即可得到亮氨酸接受能力为1400pmol/A₂₆₀单位的tRNA^Leu。     

Restoration by the rac locus of recombinant forming ability in recB? and recC? merozygotes of Escherichia coli K-12

Summary The recombinant forming ability of recB ^– or recC ^– strains of E. coli K12 is almost totally recovered in merozygotes which are heterozygous for a genetic locus denoted rac which is located five minutes clockwise from trp on the genetic map. This transient recovery phenomenon only occurs when the donor strain is rac ⁺ (wild type) and the recipient strain is rac ^-. The recombinants derived from such crosses all have the normal phenotype characteristic of recB ^– (or recC ^–) strains, and they are almost always rac ^-. The results imply that the rac ⁺ locus (or loci) is zygotically expressed and excised from the chromosome in a manner which is analogous to the zygotic induction of a prophage.     

Expression of the plasmid pKM101-determined DNA repair system in recA? and lex? strains of Escherichia coli

Summary pKM101, a plasmid R factor of the N compatibility group increases methylmethane sulfonate mutagenesis and diminishes UV-killing in recA ⁺ lex ⁺ and recA ⁺ lex ^– strains, but not in recA ^– lex ⁺ strains. The induction of a reclex dependent colicin is not present in lex ^– strains carrying the pKM101 factor. These facts indicate that pKM101 acts through an error-prone DNA repair system, which is recA ⁺ dependent, but not lex ⁺ dependent.This paper is published on the occasion of Dr. C. Callerio's seventy-fifth birthday     

Ethanol-hypersensitive and ethanol-dependent cdc? mutants in Schizosaccharomyces pombe

Ethanol-hypersensitive strains (ets mutants), unable to grow on media containing 6% ethanol, were isolated from a sample of mutagenized Schizosaccharomyces pombe wild-type cells. Genetic analysis of these ets strains demonstrated that the ets phenotype is associated with mutations in a large set of genes, including cell division cycle (cdc) genes, largely non-overlapping with the set represented by the temperature conditional method; accordingly, we isolated some ets non-ts cdc ^– mutants, which may identify novel essential genes required for regulation of the S. pombe cell cycle. Conversely, seven well characterized ts cdc ^– mutants were tested for their ethanol sensitivity; among them, cdc1–7 and cdc13–117 exhibited a tight ets phenotype. Ethanol sensitivity was also tested in strains bearing different alleles of the cdc2 gene, and we found that some of them were ets, but others were non-ets; thus, ethanol hypersensitivity is an allele-specific phenotype. Based on the single base changes found in each particular allele of the cdc2 gene, it is shown that a single amino acid substitution in the p34^cdc2 gene product can produce this ets phenotype, and that ethanol hypersensitivity is probably due to the influence of this alcohol on the secondary and/or tertiary structure of the target protein. Ethanol-dependent (etd) mutants were also identified as mutants that can only be propagated on ethanol-containing media. This novel type of conditional phenotype also covers many unrelated genes. One of these etd mutants, etd1-1, was further characterized because of the lethal cdc ^– phenotype of the mutant cells under restrictive conditions (absence of ethanol). The isolation of extragenic suppressors of etd1-1, and the complementation cloning of a DNA fragment encompassing the etd1 ⁺ wild-type gene (or an extragenic multicopy suppressor) demonstrate that current genetic techniques may be applied to mutants isolated by using ethanol as a selective agent.     

Der Bienenfresser(Merops apiaster L.) in ?sterreich

Ohne Zusammenfassung     

Red-mediated recombination of phage lambda in a recA? recB?host

Summary The lambda Red recombination system works poorly among unreplicated gam ⁺ lambda chromosomes in recA ^- cells compared to recA ⁺ cells. Recombination is not enhanced in recA ^- recB^-cells. Thus, the inability of Red to promote recombination in recA ^- replication-blocked cross is not due to the hypothetical destruction of recombination intermediates by the recB nuclease. This conclusion strengthens previous proposals that the products of the red genes can operate upon recombinational intermediates which require recA activity for their formation.     

Overexpression of Arabidopsis thaliana Na+/H+ antiporter gene enhanced salt resistance in transgenic poplar (Populus?×?euramericana ‘Neva’)

Salinity is a major abiotic stress factor limiting plant growth and productivity. One possible method to enhance plant salt-resistance is to compartmentalize sodium ions away from the cytosol. In the present work, a vacuolar Na⁺/H⁺ antiporter gene AtNHX1 from Arabidopsis thaliana, was transferred into Populus × euramericana ‘Neva’ by Agrobacterium tumefaciens in order to enhance poplar salt-resistance. The results showed that the transgenic poplar were more resistant to NaCl than the wild-type (WT) in greenhouse condition. Compared with the WT, plant growth and photosynthetic capacity of the transgenic plants were enhanced, and the transgenic plants accumulated more Na⁺ and K⁺ in roots and leaves under the same NaCl condition, whereas malondialdehyde and relative electrical conductivity were lower. All of these properties of the transgenic poplar were likely to be a consequence of the overexpression of AtNHX1 caused Na⁺ sequestration in the vacuoles and improved K⁺ absorption, thus reducing their toxic effects. These results indicated overexpression of the AtNHX1 enhanced salt-resistance of poplar, and AtNHX1 played an important role in the compartmentation of Na⁺ into the vacuoles. Therefore, this study provides an effective way for improving salt resistance in trees.     

Molecular cloning of a cysteine synthase cDNA from Citrullus vulgaris (watermelon) by genetic complementation in an Escherichia coli Cys? auxotroph

We have isolated cDNA clones encoding cysteine synthase (CSase, EC 4.2.99.8), which catalyzes the terminal step in cysteine biosynthesis, by direct genetic complementation of a Cys^– mutation in Escherichia coli with an expression library of Citrullus vulgaris (watermelon) cDNA. The library was constructed from 8-day-old etiolated seedlings of C. vulgaris in the ZAPII vector, converted to a plasmid library by in vivo excision, and then used for transformation of cysteine auxotroph E. coli NK3, which lacks the cysK and cysM loci. The complementing cDNA containing a 560 by 5-untranslated region encodes a polypeptide of 325 amino acids of M_r 34342. The translational product reacted with an antibody raised against CSase A of Spinacia oleracea. CSase and -pyrazolealanine synthase activities were demonstrated in vitro in extracts from E. coli cells expressing the cDNA. Genomic DNA blot analysis indicated the presence of a single copy of the gene, designated cysA, in the C. vulgaris genome. RNA blot hybridization indicated constitutive expression of cysA in cotyledons, hypocotyls and radicles of green and etiolated seedlings. These data suggested that this cDNA clone encodes CSase A the homolog of which in spinach is localized in the cytoplasm. The molecular phylogenetic tree of the amino acid sequences of CSaes from plants and bacteria suggested that there are three families in the CSase superfamily; the plant CSase A family, the plant CSase B family and the bacterial CSase family. The proteins in the plant CSase A family are the most conserved relative to the ancestral CSase protein.     

A ple?otropic acid phosphatase-deficient mutant of Escherichia coli shows premature termination in the dsbA gene. Use of dsbA :: phoA furions to localize a structurally important domain in DsbA

An Arabidopsis thaliana cDNA library was used to complement Saccharomyces cerevisiae pyrimidine auxotrophic mutants. Mutants in all but one (carbamylphosphate synthetase) of the six steps in the de novo pyrimidine biosynthetic pathway could be complemented. We report here the cloning, sequencing and computer analysis of two cDNAs encoding the aspartate transcarbamylase (ATCase; EC 2.1.3.2) and orotate phosphoribosyltransferase-orotidine-5-phosphate decarboxylase (OPRTase-OMP-decase; EC 2.4.2.10, EC 4.1.1.23) enzymes. These results confirm the presence in A. thaliana of a bifunctional gene whose product catalyses the last two steps of the pyrimidine biosynthetic pathway, as previously suggested by biochemical studies. The ATCase encoding cDNA sequence (PYRB gene) shows an open reading frame (ORF) of 1173 by coding for 390 amino acids. The cDNA encoding OPRTase-OMPdecase (PYRE-F gene) shows an ORF of 1431 by coding for 476 amino acids. Computer analysis of the deduced amino acid sequences of both cDNAs shows the expected high similarity with the ATCase, ornithine transcarbamylase (OTCase; EC 2.1.3.3), OPRTase and OMPdecase families. This heterospecific cloning approach increases our understanding of the genetic organization and interspecific functional conservation of the pyrimidine biosynthetic pathway and underlines its usefulness as a model for evolutionary studies.     

Acanthocephalus?amini n. sp. (Acanthocephala: Echinorhynchidae) from the freshwater fish Cichlasoma?urophthalmus (Günther) (Cichlidae) in Mexico

Acanthocephalus amini n. sp. (Palaeacanthocephala: Echinorhynchidae) is described from the intestine of Cichlasoma urophthalmus (Günther) (Pisces: Cichlidae) collected in the Río Champotón, a river in Campeche State, Mexico. It is the fourth species of Acanthocephalus Koelreuther, 1771 described from North American freshwater fishes, although two other species are known from South America. The new species is distinguished from other members of Acanthocephalus by features of its trunk, which is small, clavate, slightly expanded medially and bluntly pointed posteriorly. It is further distinguished by having a cylindrical proboscis armed with 13–14 longitudinal rows of 11–12 stout hooks; the apical and medial proboscis hooks are almost uniform in size and shape, decreasing in size towards the base; the posteriormost hooks are smaller, straighter and more slender than the anterior and middle hooks; and the lateral rows of hooks are more widely spaced, forming a conspicuous longitudinal area devoid of hooks. Furthermore, the lemnisci are saccate and shorter than the proboscis receptacle; and the neck is very short with a thick collar of trunk tegument, which encircles the base of proboscis. In males, the testes are in the middle third of trunk, diagonal, spherical and small relative to the body size, and there are six clavate cement glands. In females, the uterus forms a conspicuous, elongate, cylindrical egg reservoir. The new species is most similar to A. alabamensis Amin & Williams, 1983, but can be distinguished by its swollen, clavate trunk, the largest proboscis hooks being present apically and medially, smaller testes, a shorter male reproductive system relative to body size and females with a prominent uterus. They have different hosts and geographical distribution. The new species can be differentiated from Brasacanthus sphoeroides Thatcher, 2001, a similar species in a monotypic echinorhynchid genus, because the latter is larger, has smaller proboscis hooks and its lemnisci are longer than the receptacle.     

Isolation of Rec? mutants from an F-prime merodiploid strain of Escherichia coli K-12

Summary This paper describes a method of screening mutagenised populations of an E. coli gal _A ^– gal _B ^– F-prime merodiploid for mutants defective in recombination. The method relies on scoring colonies on Eosin-Methylene Blue agar that have fewer than normal numbers of Gal⁺ papillae. With a suitable choice of gal ^–mutations most of the papillae arise by recombination and some of those colonies with less than normal numbers prove to be defective in some aspect of recombination or DNA repair. In addition to strains carrying mutations that can be ascribed to known loci, several novel mutant phenotypes were identified.     

Role of quinones in electron transport to oxygen and nitrate in Escherichia coli. Studies with a ubiA− menA− double quinone mutant

A ubiA^? menA^? double quinone mutant of Escherichia coli K12 was constructed together with other isogenic strains lacking either ubiquinone or menaquinone. These strains were used to study the role of quinones in electron transport to oxygen and nitrate. Each of the four oxidases examined (NADH, d-lactate, α-glycerophosphate and succinate) required a quinone for activity. Ubiquinone was active in each oxidase system while menaquinone gave full activity in α-glycerophosphate oxidase, partial activity in d-lactate oxidase but was inactive in NADH and succinate oxidation. The aerobic growth rates, growth yields and products of glucose metabolism of the quinone-deficient strains were also examined. The growth rate and growth yield of the ubi⁺ menA^? strain was the same as the wild-type strain, whereas the ubiA^? men⁺ strain grew more slowly on glucose, had a lower growth yield (30% of wild type) and accumulated relatively large quantities of acetate and lactate. The growth of the ubiA^? menA^? strain was even more severely affected than that of the ubiA^? men⁺ strain.Electron transport from formate, d-lactate, α-glycerophosphate and NADH to nitrate was also highly dependent on the presence of a quinone. Either ubiquinone or menaquinone was active in electron transport from formate and the activity of the quinones in electron transport from the other substrates was the same as for the oxidase systems. In contrast, quinones were not obligatory carriers in the anaerobic formate hydrogenlyase system. It is concluded that the quinones serve to link the various dehydrogenases with the terminal electron transport systems to oxygen and nitrate and that the dehydrogenases possess a degree of selectivity with respect to the quinone acceptors.     

Measurements of the gas temperature in an oxygen plasma by spectroscopy of the O2( b 1Σ g+ ) → O2( X 3Σ g? ) transition

A method for measuring the gas temperature in an oxygen plasma by spectroscopy of the electronic transition from the O₂(b ¹Σ _g ⁺ , v = 0) metastable state of molecular oxygen into the O₂(X ³Σ _g ⁻ , v = 0) ground state is considered in detail. The method is verified experimentally for the plasma of dc glow discharge in pure oxygen. It is shown that the gas temperature can be determined by analyzing high-resolution spectra of the P branch of this transition, no matter whether its fine structure (^P P and ^P Q branches) is resolved or masked, provided that the rotational structure of the spectrum is resolved. The feasibility of the method proposed in 1999 by P. Maco and P. Veis for determining the gas temperature from the ratio between the intensity maxima of the R and P branches of the O₂(b ¹Σ _g ⁺ , v = 0) → O₂(X ³Σ _g ⁻ , v = 0) transition in a poorly resolved spectrum was studied experimentally. It is shown that, in order to use this method, it is necessary to know the spectrograph instrumental function. The effect of the spatial inhomogeneity of the temperature and concentration of O₂(b ¹Σ _g ⁺ ) molecules on the accuracy of integral (over the plasma volume) measurements of the gas temperature is investigated using spatially resolved spectroscopy of the O₂(b ¹Σ _g ⁺ , v = 0) → O₂(X ³Σ _g ⁻ , v = 0) transition. It is shown that precise measurements of the temperature require that the optical measurement system be thoroughly adjusted in order for the temperature and concentration of the emitting particles to vary insignificantly over the optically selected volume. Original Russian Text ? S.M. Zyryanov, D.V. Lopaev, 2007, published in Fizika Plazmy, 2007, Vol. 33, No. 6, pp. 563–574.     

A ThCAP gene from Tamarix hispida confers cold tolerance in transgenic Populus (P. davidiana?×?P. bolleana)

The ThCAP gene, which encodes a cold acclimation protein, was isolated from a Tamarix hispida NaCl-stress root cDNA library; its expression patterns were then assayed by qRT-PCR in different T. hispida tissues treated with low temperature (4°C), salt (400 mM NaCl), drought (20% PEG6000) and exogenous abscisic acid (100 μM). Induction of ThCAP gene was not only responsive to different stress conditions but was also organ specific. When transgenic Populus (P. davidiana × P. bolleana) plants were generated, expressing ThCAP under regulation of the cauliflower mosaic virus CaMV 35S promoter, they had a greater resistance to low temperature than non-transgenic seedlings, suggesting that ThCAP might play an important role in cold tolerance.     

Eine mendelnde, k?lteempfindliche Sippe(f. delicata) derMirabilis Jalapa

Ohne ZusammenfassungMit 1 Abbildung im Text.     

Over-expression of the Arabidopsis Na+/H+ antiporter gene in Populus deltoides CL?×?P. euramericana CL “NL895” enhances its salt tolerance

Soil salinity is a serious problem worldwide. It is necessary to improve the salt tolerance of plants to avoid the progressive deterioration of saline soil. We showed that the over-expression of AtNHX1 improves salt tolerance in a transgenic poplar (Populus deltoides CL × P. euramericana CL “NL895”) under mannose selection. Four transgenic poplar plants were obtained. Southern blot analysis showed that the pmi gene had integrated into the genome of the poplar. RT-PCR confirmed that AtNHX1 could be expressed normally in the transgenic plants. When tested for salt tolerance by NaCl stress, we measured a 100% increase in Na⁺ content in the three transgenic lines (T18, T50, T98) significantly higher than the 33% increase seen in wild-type plants. The chlorophyll content of the transgenic plants was not altered significantly, while the chlorophyll content in the control plants showed a small decrease. MDA content was decreased in the transgenic plants. These results show that the AtNHX1 gene may enhance salt tolerance due to increased vacuolar compartmentalization of sodium ions.     

Escherichia coli K-12 F? mutants that form mating aggregates but form transconjugants with low frequencies

Summary We have isolated Escherichia coli F^– mutants which, when mated with either Hfr or F, can form stable mating aggregates well but produce transconjugants with reduced frequencies. Selection procedure and other tests rule out the possibility that they are Rec^– strains. These mutants can be classified into two types: type I mutants can induce conjugal DNA replication in the donor, yet form transconjugants poorly; whereas, type II mutants induce conjugal DNA replication with poor efficiencies in the donor. Further tests indicate that type I mutants are very sensitive to lethal zygosis and their membranes, both inner and outer, show alterations in protein composition, whereas type II mutants are insensitive to lethal zygosis, and have an obvious alteration in the protein composition of their outer membrane. These results suggest that type I is defective in transconjugant formation primarily due to a change in the inner membrane, whereas type II is defective in generating a mating signal, which induces donor conjugal DNA replication, due to an alteration in the outer membrane.     

DnaK-dependent ribosome biogenesis in Escherichia coli? : competition for dominance between the alleles dnaK756 and dnaK ?+

The Escherichia coli chaperone DnaK is vital for many cellular functions, including ribosome biogenesis at high temperature. Thus, the dnaK756-ts (λ ^R ) mutant, at the non-permissive temperature, is inhibited at a late stage of ribosome assembly, yielding 21S, 32S and 45S precursor particles. This defect, unlike the λ resistance and thermosensitivity phenotypes, is not complemented by lysogenisation with a transducing phage λ dnaK ⁺ bearing the wild-type dnaK gene. However this dominant phenotype becomes recessive when dnaK ⁺ is expressed from a medium-copy-number plasmid. On the other hand, an excess of DnaK causes an unexpected dominant-lethal effect of the dnaK756 allele near non-permissive temperatures. This interplay between the dnaK ⁺ and dnaK756 alleles supports the idea of that DnaK oligomers form in the cell. Received: 28 April 1998 / Accepted: 24 July 1998     

Circaetus gallicus (Gm) in Holstein?

Ohne Zusammenfassung