Vacuum blotting enhances nucleic acid transfer

Nucleic acids can be transferred from an agarose get to a transfer membrane using capillary Southern blotting or electroblotting techniques. Vacuum blotting represents an alternative method that offers a number of advantages.     

Expanding nucleic acid function in vitro and in vivo

Nucleic Acids composed of the five natural bases and a phosphate backbone can be designed or evolved to have a wide variety of sequence-dependent functions. Recent in vitro work has addressed some outstanding issues in evolving nucleic acid catalysts, as well as the creation of prescribed shapes and arrays from oligonucleotides and long single-stranded nucleic acids. Nucleic acids have also been engineered in vivo, leading to new modes of gene regulation. It is likely that the improving ability to synthesize long DNA sequences will accelerate the creation of novel functions from nucleic acids.     

Isolation of deoxyribonucleic acid and ribosomal ribonucleic acid from bacteria

1. Nucleic acids were released from Escherichia coli by lysing with tri-iso-propylnaphthalene sulphonate and 4-aminosalicylate and then extracting with a phenol-cresol mixture. 2. Nucleic acids were similarly released from Bacillus subtilis after initial treatment with lysozyme. 3. DNA was sedimented after careful precipitation with m-cresol or 2-butoxyethanol (0.1-0.12vol.) in the presence of 20% sodium benzoate. 4. Contaminating ribosomal RNA was removed by precipitation in the presence of 4m-sodium chloride or by extracting DNA with an acetate-butyrate mixture, in which RNA is insoluble. 5. The DNA from B. subtilis has a transforming ability of 0.3-0.6% for the tryptophan marker. 6. Ribosomal RNA was then precipitated with rapidly labelled RNA by the addition of an equal volume of 2-butoxyethanol. 7. There was good separation of the nucleic acids from protein and polysaccharides.     

日本稻蝗、中华稻蝗和赤胫伪稻蝗地理种群的RAPD遗传分化研究

运用10个RAPD引物对日本稻蝗(Oxya japonica)、中华稻蝗(Oxya chinensis)和赤胫伪稻蝗(Pseudoxya diminuta)的种群遗传分化进行分析。10个随机引物共产生135条带,扩增谱带具有明显的属、种间多态性。Shannon信息指数表明中华稻蝗遗传多样性水平较高(2．693),日本稻蝗次之(2.319),赤胫伪稻蝗最低(1．042)。中华稻蝗和日本稻蝗的不同地理居群出现遗传分化,由Shannon信息指数估算的种群间遗传分化系数分别为20．7％,42．4％。用UPGMA和NJ法对Nei’s遗传距离作聚类分析,构建分子系统树。系统树显示：同一种群的不同个体优先相聚,而后,同一种的不同种群依次相聚;日本稻蝗广西南宁种群和广东广州种群首先聚为一支,陕西西安种群和浙江杭州种群聚为另一支,两支相聚后与中华稻蝗聚在一起,最后与赤胫伪稻蝗相聚。聚类结果表明：不同地域日本稻蝗亲缘关系的远近与地理距离呈现一定的相关趋势,日本稻蝗与中华稻蝗亲缘关系较近,Nei’S遗传距离平均为0．142,而赤胫伪稻蝗与它们关系较远,Nei’s遗传距离平均为0．451。聚类图所显示的物种间亲缘关系的远近程度与形态分类学和细胞分类学结果相一致。     

Detection of target DNA using fluorescent cationic polymer and peptide nucleic acid probes on solid support

Background  

Nucleic acids detection using microarrays requires labelling of target nucleic acids with fluorophores or other reporter molecules prior to hybridization.     

Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent

Nucleic acid can catalyze the conversion of α‐helical cellular prion protein to β‐sheet rich Proteinase K resistant prion protein oligomers and amyloid polymers in vitro and in solution. Because unfolding of a protein molecule from its ordered α‐helical structure is considered to be a necessary step for the structural conversion to its β‐sheet rich isoform, we have studied the unfolding of the α‐helical globular 121–231 fragment of mouse recombinant prion protein in the presence of different nucleic acids at neutral and acid pH. Nucleic acids, either single or double stranded, do not have any significant effect on the secondary structure of the protein fragment at neutral pH; however the protein secondary structure is modified by the nucleic acids at pH 5. Nucleic acids do not show any significant effect on the temperature induced unfolding of the globular prion protein domain at neutral pH which, however, undergoes a gross conformational change at pH 5 as evidenced from the lowering of the midpoint of thermal denaturation temperatures, Tm, of the protein. The extent of Tm decrease shows a dependence on the nature of nucleic acid. The interaction of nucleic acid with the nonpolar groups exposed from the protein interior at pH 5 probably contributes substantially to the unfolding process of the protein.     

乳酸杆菌发酵滤液中核酸抗肿瘤效应及对荷瘤鼠免疫功能的影响

目的探讨乳酸杆菌DM9811培养基滤液中核酸在体外对肿瘤细胞生长及对荷瘤鼠免疫功能的影响。方法抽提乳酸杆菌DM9811对数生长期培养基滤液中核酸,进行琼脂糖电泳分析;MTT法体外观察其对于结肠癌HT-29细胞系生长的影响;动物实验观察不同组别小鼠生存期、瘤重体重比、NK细胞活性、T细胞亚群指标。结果乳酸杆菌DM9811滤液中核酸组分为RNA;在细胞浓度为1×106时,每孔加入RNA 100μg能够明显抑制结肠癌HT-29细胞系的生长;预防组荷瘤鼠除CD8＋T细胞比例之外各项指标与阴性对照组相比差异都有统计学意义（P〈0.05）,其中NK细胞杀伤活性（58.97±3.62）、CD4＋T细胞比例（27.77±5.40）和生存期（15.1±4.48）均高于阴性对照组（43.87±3.92）、（19.68±3.00）、（10.2±3.08）,瘤重体重比（0.029±0.017）低于阴性对照组（0.066±0.024）;治疗组生存期（11.8±3.12）与阴性对照组（10.2±3.08）相比差异有统计学意义（P〈0.05）,治疗组生存期长于阴性对照组。结论乳酸杆菌DM9811培养基滤液中存在核酸组分,为100200 bp的RNA片段。100μg/mL乳酸杆菌DM9811培养基滤液中RNA组分对结肠癌HT-29细胞系生长有明显抑制作用。RNA组分可以上调荷瘤鼠的细胞免疫水平,延长荷瘤鼠的生存期。     

乳杆菌对数生长期培养基滤液核酸组分分析

目的通过对乳杆菌对数生长期培养基滤液中核酸组分的分析,阐明乳杆菌DM9811对数生长期培养基滤液中核酸的性质。方法应用核酸的分离、纯化及电泳分析技术。结果乳杆菌对数生长期培养基滤液中核酸组分为RNA,其片段大小为100 bp左右。乳杆菌对数生长期培养基滤液中核酸组分为RNA,为对数期产生且呈时间依赖关系。结论核酸组分不仅仅是遗传信息的载体,还可能作为有效的信息分子。     

An ethidium-acrylamide affinity medium for recovery of nucleic acids from free solution and from polyacrylamide and agarose gels

The simple preparation of an ethidium-bromide-based nucleic acid affinity medium is described. The medium is composed of an acrylamide matrix to which ethidium bromide is attached. Its use in preparative purification and fractionation of nucleic acids in solution and in electrophoretic elution of nucleic acids from gels is reported. Nucleic acids can be eluted from this medium with a buffered salt solution and concentrated by ethanol precipitation without persistent contamination with undesirable impurities.     

Modified Oligonucleotides: New Structures,New Properties,and New Spheres of Application

Russian Journal of Bioorganic Chemistry - Nucleic acids have made a long and arduous journey “from the bench to the bedside.” At present, it can be assumed that drugs based on...     

Building objects from nucleic acids for a nanometer world

Nucleic acids are an ideal material for the construction of nanometer-scaled objects. An overview is given which focuses on the structural aspects of this field of research using native DNA and RNA and especially also chemically modified derivatives, which offer structural elements other than the Watson-Crick interaction. First examples for applications are discussed.     

Exploring the impact of nucleic acids on protein stability in bacterial cell lysate

- Addition of salt enhanced thermal stability of model substrate proteins by reducing electrostatic repulsion between protein molecules.- However, the opposite effect was observed with bacterial cell lysate, indicating that certain molecules within the lysate could enhance protein stability via electrostatic interactions.- Such molecules present in cell lysate were found to be nucleic acids known to have a potent anti-aggregation activity toward proteins involving electrostatic interactions.- Nucleic acids showed chaperone activity in physiological salt concentration within cells and in buffer or medium commonly used in experiments.- The chaperone activity of nucleic acids should be taken into account when performing various in vitro assays using cell lysate or samples containing nucleic acids.     

Electrophoretic elution of nucleic acids from acrylamide and agarose gels

A simple method for electrophoretic elution of nucleic acids from gel slices is described. The procedure utilizes a standard tube gel system and can be completed in as little as one hour. Nucleic acids are recovered in a small volume with almost 100% efficiency. The procedure is applicable equally to acrylamide and agarose gels, and small as well as large RNA and DNA molecules. The eluted nucleic acids are essentially undegraded and are suitable for a variety of structural and biological analyses.     

Binding and purification of plasmid DNA using multi-layered carbon nanotubes

We propose a new method for the separation of nucleic acids using multi-layered carbon nanotubes (CNTs) as an adsorbent. According to agarose gel electrophoresis, oxidized water-stable CNTs adsorb certain forms of nucleic acids, such as high molecular weight RNA, chromosomal DNA, linear and denatured forms of plasmid DNA. However, CNTs do not adsorb supercoiled form of plasmid DNA. Nucleic acids bound to CNTs can be readily removed by centrifugation whereas supercoiled plasmid DNA remains in solution. Upon the addition of divalent metal ions supercoiled plasmid DNA forms relatively stable complexes with CNTs due to chelation. Thus, new details about association of nucleic acids with CNTs were revealed and stoichiometry of the complexes was estimated. Our results can be used for fine purification of supercoiled plasmid DNA for gene therapy applications as well as manipulation of nucleic acids for biosensor design.     

Fungal nucleic acids as interferon inducers.

Nucleic acids isolated from the fungi Aspergillus niger x11, Piptoporus betulinus and Ganoderma applanatum reduced the number of vaccinia virus plaques in chick embryo fibroblast (CEF) tissue culture and when administered intravenously to white mice protected them against lethal infection with tick borne encephalitis virus strain K5 (TBE). In CEF tissue culture the nucleic acids of the studied fungi were found to induce small but detectable amounts of a substance with the character of interferon. In vivo only ribonucleic acid from G. applanatum induced a substance showing interferon properties in the spleen of mice.     

Buoyant density sedimentation of macromolecules in sodium iothalamate density gradients.

Nucleic acids, bacteriophages, phage capsids, and a DNA-capsid complex have been centrifuged to an equilibrium buoyant density in sodium iothalamate density gradients. Nucleic acids have comparatively high hydrations and are less dense than proteins in these gradients. Sodium iothalamate gradients can be used to separate DNA from RNA, single-chain DNA from double-chain DNA and to separate bacteriophage T7 and λ deletion mutants from the respective wild-type phage.The DNA packaged in bacteriophage T7 appears to be less hydrated than free DNA in sodium iothalamate gradients. There is evidence that the hydration of DNA packaged in phage T7 is restricted by the volume of the phage head. The total volume of phage T7 was estimated to be 1.32 × 10⁻¹⁶ ml. The volume available to package phage T7 DNA was estimated to be 2.2 times the volume of the B form of T7 DNA.     

Nucleic acids in mummified plant seeds: biochemistry and molecular genetics of pre-Columbian maize.

Nucleic acids fractions were isolated from pre-Columbian maize seeds and characterized using different approaches such as polyacrylamide gel electrophoresis, anti-DNA antibody binding, HPLC fractionation, molecular hybridization with cloned genes, and DNA amplification by the polymerase chain reaction. The nucleic acids were found to be very depolymerized (less than or equal to 140 base pairs in length) and composed mainly of ribosomal RNA. Despite the very low amount and degree of polymerization of seed DNA, specific maize nuclear Mu1, Mu4, Mu8 and, possibly, Mu5 element components could be detected, thanks to the use of amplification systems as short as 90 bp. The results suggest that evaluation of the relative proportions of Mu-type element components and, possibly, other maize genomic components in single mummified kernels, may offer a new key to the study of ancient maize populations.     

Anion binding to nucleic acids

Nucleic acids are generally considered as efficient cation binders. Therefore, the likelihood that negatively charged ions might intrude their first hydration shell is rarely considered. Here, we show on the basis of (i) a survey of the Nucleic Acid Database, (ii) several structures extracted from the Cambridge Structural Database, and (iii) molecular dynamics simulations, that the nucleotide electropositive edges involving mainly amino, imino, and hydroxyl groups can cast specific anion binding sites. These binding sites constitute also good locations for the binding of the negatively charged groups of the Asp and Glu residues or the nucleic acid phosphate groups. Furthermore, it is observed in several instances that anions, like water molecules and cations, do mediate protein/nucleic acid interactions. Thus, anions as well as negatively charged groups are directly involved in specific recognition and folding phenomena involving polyanionic nucleic acids.     

Activity of glucose-6-phosphate dehydrogenase and level of nucleic acids in Escherichia coli during various rates of growth]

Nucleic acids content and activity of glucose-6-phosphate dehydrogenase were studied at various growth rates. The activity of glucose-6-phosphate dehydrogenase was shown to correlate with the growth rate and with the DNA content in Escherichia coli.