Tn10 transposition via a DNA hairpin intermediate

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder characterized by microcephaly, a birdlike face, growth retardation, immunodeficiency, lack of secondary sex characteristics in females, and increased incidence of lymphoid cancers. NBS cells display a phenotype similar to that of cells from ataxia-telangiectasia patients, including chromosomal instability, radiation sensitivity, and aberrant cell-cycle-checkpoint control following exposure to ionizing radiation. A recent study reported genetic linkage of NBS to human chromosome 8q21, with strong linkage disequilibrium detected at marker D8S1811 in eastern European NBS families. We collected a geographically diverse group of NBS families and tested them for linkage, using an expanded panel of markers at 8q21. In this article, we report linkage of NBS to 8q21 in 6/7 of these families, with a maximum LOD score of 3.58. Significant linkage disequilibrium was detected for 8/13 markers tested in the 8q21 region, including D8S1811. In order to further localize the gene for NBS, we generated a radiation-hybrid map of markers at 8q21 and constructed haplotypes based on this map. Examination of disease haplotypes segregating in 11 NBS pedigrees revealed recombination events that place the NBS gene between D8S1757 and D8S270. A common founder haplotype was present on 15/18 disease chromosomes from 9/11 NBS families. Inferred (ancestral) recombination events involving this common haplotype suggest that NBS can be localized further, to an interval flanked by markers D8S273 and D8S88.     

Hairpin-dimer equilibrium of a parallel-stranded DNA hairpin: formation of a four-stranded complex

The 24mer deoxyoligonucleotide 3'-d(T)10-5'-5'-d(C)4- d(A)10-3'(psC4) with an uncommon 5'-p-5'phosphodiester linkage was designed to enable the formation of a hairpin structure with unusual parallel-stranded stem. As reference hairpin structure with an antiparallel-stranded stem, the 24mer 5'-d(T)10-d(C)4-d(A)10-3'(apsC4) was chosen. The behaviour of these oligonucleotides at different temperatures, DNA and salt concentrations was characterised by a combination of UV melting, CD, CD melting, infrared and Raman spectroscopy, infrared melting and analytical ultracentrifugation. The parallel-stranded hairpin structure was found to be formed by psC4 only under conditions of low DNA concentration and low salt concentration. Increase of the NaCl concentration beyond the physiological level or high DNA concentration supports the formation of intermolecular multi-stranded structures. The experimental data are in agreement with a four-stranded complex formed by two molecules of psC4. The base pairing model of this asymmetric four-stranded complex is based on the pyrimidine motif of a triple helix with two bifurcated hydrogen bonds at the O4 of the thymine each directed towards one of the amino protons of both adenines. In contrast, the reference oligonucleotide apsC4 forms only an antiparallel-stranded hairpin under all experimental conditions.     

Three-dimensional model of a hairpin ribozyme

On the basis of the mutational and chemical-modification analysis, we propose the new secondary structure of a hairpin ribozyme, which contains three stems, a triple helix and the reverse Watson-Crick g+1C44 base pair at the 3' side of the cleavage site. The computational approach to the tertiary structure of a hairpin ribozyme have been carried out.     

等温熔炼炉开创铝熔炼技术新时代

节约两源（资源与能源）与减少污染物排放是一个永恒的话题。铝工业特别是原铝生产工业既是高能耗工业又是高污染行业，节能减排有着重要意义。铝工业下游产品行业的单位产品平均能耗虽然还不到上游产品的1/18，但由于产量大，总能耗高，也应是节能重点行业之一。由美国能源部拨款，阿波格技术公司、爱励公司、德雷克塞尔大学、阿贡国家实验室联合研发的铝合金等温熔炼炉（ITM）在铝及铝合金熔炼技术发展史上有着划时代的意义，与当前的常规反射炉相比，可节能70％，减排80％，铝的烧损下降4个百分点，在节源（资源与能源）减排方面意义重大。等温熔炼炉在美国已商业化试生产成功，正在作扩大试生产工作，并开发大的商业化生产炉，预计2010年美国可能有20％左右的下游铝产品是用此项技术熔炼的，首先得到应用的是铸造行业与压铸件生产行业。此技术不但适合于铝工业，也适用于其他冶金部门，还可用于熔化玻璃。     

Model Experiments on Melting of Alloys with Low Melting Temperature in a Medium with High Melting Temperature

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High-mobility group 1/2 proteins are essential for initiating rolling-circle-type DNA replication at a parvovirus hairpin origin

Rolling-circle replication is initiated by a replicon-encoded endonuclease which introduces a single-strand nick into specific origin sequences, becoming covalently attached to the 5' end of the DNA at the nick and providing a 3' hydroxyl to prime unidirectional, leading-strand synthesis. Parvoviruses, such as minute virus of mice (MVM), have adapted this mechanism to amplify their linear single-stranded genomes by using hairpin telomeres which sequentially unfold and refold to shuttle the replication fork back and forth along the genome, creating a continuous, multimeric DNA strand. The viral initiator protein, NS1, then excises individual genomes from this continuum by nicking and reinitiating synthesis at specific origins present within the hairpin sequences. Using in vitro assays to study ATP-dependent initiation within the right-hand (5') MVM hairpin, we have characterized a HeLa cell factor which is absolutely required to allow NS1 to nick this origin. Unlike parvovirus initiation factor (PIF), the cellular complex which activates NS1 endonuclease activity at the left-hand (3') viral origin, the host factor which activates the right-hand hairpin elutes from phosphocellulose in high salt, has a molecular mass of around 25 kDa, and appears to bind preferentially to structured DNA, suggesting that it might be a member of the high-mobility group 1/2 (HMG1/2) protein family. This prediction was confirmed by showing that purified calf thymus HMG1 and recombinant human HMG1 or murine HMG2 could each substitute for the HeLa factor, activating the NS1 endonuclease in an origin-specific nicking reaction.     

Structural features of the DNA hairpin d(ATCCTA-GTTA-TAGGAT): formation of a G-A base pair in the loop

The three-dimensional structure of the hairpin formed by d(ATCCTA-GTTA-TAGGAT) has been determined by means of two-dimensional NMR studies, distance geometry and molecular dynamics calculations. The first and the last residues of the tetraloop of this hairpin form a sheared G-A base pair on top of the six Watson-Crick base pairs in the stem. The glycosidic torsion angles of the guanine and adenine residues in the G-A base pair reside in the anti and high- anti domain ( approximately -60 degrees ) respectively. Several dihedral angles in the loop adopt non-standard values to accommodate this base pair. The first and second residue in the loop are stacked in a more or less normal helical fashion; the fourth loop residue also stacks upon the stem, while the third residue is directed away from the loop region. The loop structure can be classified as a so-called type-I loop, in which the bases at the 5'-end of the loop stack in a continuous fashion. In this situation, loop stability is unlikely to depend heavily on the nature of the unpaired bases in the loop. Moreover, the present study indicates that the influence of the polarity of a closing A.T pair is much less significant than that of a closing C.G base pair.     

Selective photocleavage of DNA by anthraquinone derivatives: targeting the single-strand region of hairpin structures

A tetracationic anthraquinone derivative (27AQS2) binds to hairpin DNA and irradiation of the bound quinone leads to selective strand cleavage. NMR spectroscopy reveals that 27AQS2 binds at the loop and to the stem-loop junction of hairpin DNA. UV irradiation of the bound quinone causes cleavage of the DNA in the loop region and at guanines in the stem region. Inclusion of ethidium bromide in the reaction mixture leads to a greatly increased selectivity for loop cleavage. Spectroscopic and chemical evidence suggests a three component mechanism for reaction. The ability to target single-stranded regions of DNA structures is an important property of this photonuclease.     

The SbcCD nuclease of Escherichia coli is a structural maintenance of chromosomes (SMC) family protein that cleaves hairpin DNA

Hairpin structures can inhibit DNA replication and are intermediates in certain recombination reactions. We have shown that the purified SbcCD protein of Escherichia coli cleaves a DNA hairpin. This cleavage does not require the presence of a free (3' or 5') DNA end and generates products with 3'-hydroxyl and 5'-phosphate termini. Electron microscopy of SbcCD has revealed the "head-rod-tail" structure predicted for the SMC (structural maintenance of chromosomes) family of proteins, of which SbcC is a member. This work provides evidence consistent with the proposal that SbcCD cleaves hairpin structures that halt the progress of the replication fork, allowing homologous recombination to restore DNA replication.     

P nucleotides, hairpin DNA and V(D)J joining: making the connection

The fine-structures of V(D)J joining products present a cache of information about the recombination process. Testable hypotheses with predictive value are beginning to provide a surprisingly detailed picture of the joining operation. Here, one feature of V(D)J junctions, the appearance of short palindromic inserts, is discussed, and experimental evidence in support of a joining model involving a hairpin-terminated DNA intermediate will be summarized.     

无筛板沸腾氯化与熔盐氯化生产TiCl4工艺浅析

无筛板沸腾氯化法与熔盐氯化法是目前生产TiCl4的两种主要工艺,分别是我国和前苏联钛冶金工作者针对各自地区所拥有的可用于TiCl4生产的原料而独创的生产方法,均对世界钛冶金工业的发展做出了巨大贡献。首先简要概述了两种氯化法生产TiCl4的特点,对比分析了两种方法每生产1t粗TiCl4所消耗的原料和产生的废料,以及各自的优缺点,指出采用无筛板沸腾氯化法生产的粗TiCl4的质量不如熔盐氯化法,我国的无筛板沸腾氯化法仍需继续改进。     

Chemical synthesis of H-phosphonate DNA without using N-protecting groups

Oligodeoxyribonucleotides were synthesized by using N-unprotected H-phosphonate monomers. It was found that the amino groups of nucleosides were not modified during condensation where benzotriazolyloxy carbonium and phosphonium types of compounds were employed as condensing reagents. The most effective condensing reagent for rapid internucleotidic bond formation was found to be 2-(benzotriazol-1-yloxy)-1,1-dimethyl-2-(pyrrolidin-1- yl)-1,3,2- diazaphospholidinium hexafluorophosphate (BOMP). In the present H-phosphonate approach, 2-benzenesulfonyl-3-(3-nitrophenyl)oxaziridine (BNO) was successfully employed as a new oxidizing reagent for oxidation of the H-phosphonate linkages under anhydrous conditions.     

Thermodynamic analysis of an RNA combinatorial library contained in a short hairpin

Prediction of nucleic acid structure from sequence requires thermodynamic parameters for a variety of motifs, many of which are complex and consist of a large number of possible sequence combinations. Here we report an experimental approach for identifying the stable and unstable members of an RNA combinatorial library. Short model RNA hairpins consisting of 13 base pairs (bp) flanked by primer binding sites are constructed and separated according to their relative thermodynamic stabilities using temperature gradient gel electrophoresis (TGGE). Partially denaturing TGGE is carried out with potassium chloride, sodium chloride, or magnesium chloride salts in the gel. The TMs of model hairpins can be tuned by adjusting the concentration of urea in the gel while maintaining the correct order of stabilities for the hairpins. Mixtures of RNAs differing by a single Watson-Crick base pair are resolved according to their relative thermodynamic stabilities, as are mixtures of GC or AU base pair transversions differing in DeltaG degrees37 by only 0.3-0.5 kcal/mol. In addition, a simple combinatorial library with one position of randomization opposite a guanosine is prepared and separated into its four members by parallel and perpendicular TGGE. The order of thermodynamic stabilities for the library determined by TGGE is shown to be the same when assayed by UV-melting experiments. Analysis of the thermodynamics of folding of combinatorial libraries is general and may be applied to a wide variety of complex nucleic acid secondary and tertiary motifs in order to identify the stable and unstable members.