清除蚊子的两种方法

蚊子会侵扰民众,尤其是农村更为突出,同时它是传播疾病的“帮凶”。清除蚊子有多条途径,其中有两种方法值得注意：     

微生物气溶胶检测技术的研究进展

生物气溶胶中微生物的种类复杂多样,其采集和鉴定是全面掌握微生物气溶胶生物特性的关键,对防控气溶胶病原体传播十分重要.本文简要综述了微生物气溶胶的生物特性和潜在危害性,介绍了微生物气溶胶的一般采集方法,采集器采集生物气溶胶的基本原理、特点和优缺点.将有助于研究人员根据实验目的在采集低浓度生物气溶胶时选择合适的采样器.本文...     

Mu外翻的生物力学研究进展

Mu外翻是足母趾向外侧过度倾斜所引起的一种足的畸形,会产生疼痛,对人的行走功能有很大影响,近年在临床骨科得到越来越多的重视,陈宝兴统计治疗方法将近200余种。用生物力学方法研究Mu外翻引起的足的变化,有助于支足母外翻的进一步认识和对治疗的改进。     

基因治疗中关于非病毒载体的最新设计策略

基因转移的非病毒技术近来发展迅速。与病毒转染细胞的方法相比,非病毒转移方法比较简便,安全,毒性小。这种方法大体可分为两种：完全非病毒方法和病毒增强的转移方式。本文主要介绍一些最近两三年来新兴的技术方案的优点和不足。     

An improved culture system for virus isolation and detection

Cell culture plays an important role in virology. It provides a platform for the detection and isolation of viruses as well as for the biochemistry and molecular biology based studies of viruses. In the present work, a new system that could permits multiple (different) cell lines to be simultaneously cultured in one dish was developed. In the system, each cell line was cultured in an isolated zone in the same dish or well and the system is therefore called an isolated co-culture system. The usefulness of this novel approach for virus isolation was demonstrated using a model system based on adenovirus.     

口吃的症状分类

难发性：第一个字的发音特别困难，尽管做了很大的努力，也只能在喉部发出低微、沉闷的声音，如“嗯……嗯……嗯……大家好”。     

供水及间甲酚对小麦间作蚕豆土壤微生物多样性和酶活性的影响

通过盆栽试验,探讨供水(田间持水量的45%、60%和75%)和化感物质间甲酚对小麦、蚕豆不同种植模式生长盛期土壤微生物多样性和酶活性的影响.结果表明,随灌水水平的降低,不同处理的土壤细菌、真菌和放线菌数量随之减少,间甲酚可加剧灌水减少引起的微生物数量的减少;间甲酚对不同处理土壤微生物多样性指数均具有降低作用,提高灌水水平可缓解间甲酚对间作群体土壤微生物多样性的负效应,但间甲酚在75%灌水水平下对单作微生物多样性的负效应最大,45%的供水水平和间甲酚作用下间作可维持更高的土壤微生物多样性.间甲酚对土壤过氧化氢酶的化感作用不显著,对脲酶和酸性磷酸酶活性的化感作用显著;3种土壤酶活性随供水水平的降低均显著下降,但供水与间甲酚、种植模式的互作效应对酶活性的影响不显著;间作对土壤过氧化氢酶和酸性磷酸酶活性具有极显著影响.     

对硫酸软骨素传统提取方法的改进

软骨中硫酸软骨素与蛋白质结合成蛋白多糖,并与胶原蛋白结合在一起。本文采用先高温蒸煮,后加稀碱与酶解相结合提取该药物,ＴＣＡ沉淀蛋白质后,高岭土吸附,再用氯仿连续反萃取,使产品质量达到优级纯,较其他方法缩短了原工艺流程,提高了纯度,减轻了碱－盐提取所带来的环境污染。     

研究植物根系分泌物的方法

文章对植物根系分泌物的定义、组成、分类、影响因素和分泌部位,根系分泌物的收集、分离及鉴定方法进行了概述,并对此问题的今后研究方向作了展望。     

巴东木莲核基因组微卫星位点分离的初步研究

FIASCO是一种高效构建微卫星富集文库的方法。本研究采用FIASCO（fast isolation by AFLP sequences containing repeats）方法成功构建了巴东木莲（Manglietia patungensis）微卫星AC富集文库。我们对AC富集文库中的119个阳性克隆进行测序,其中57个含有微卫星序列。设计并合成了其中40对微卫星引物进行PCR扩增检测,有6对引物扩增出目的片段,然而没有位点都呈现多态性。最后对巴东木莲微卫星位点的分离效率低下的原因进行了初步探讨。     

磁珠富集法筛选大弹涂鱼CA微卫星序列

以大弹涂鱼(Boleophthalmus pectinirostris)基因组DNApool为材料,构建基因组PCR文库,采用链霉亲和素包被的磁珠富集法筛选目的片段.目的片段经克隆、测序验证后获得大弹涂鱼微卫星序列.在筛选的409个菌落中共获得209个阳性克隆,其中具有144个微卫星序列(GenBank登录号为HQ852252 - HQ852395),除去重复测序和侧翼链不足的序列,可以设计引物的微卫星序列有117条.     

黑斑狗鱼部分基因组文库构建和微卫星位点的筛选

采用磁珠富集与放射性杂交相结合的方法开发黑斑狗鱼(Esoxreieherti Dybowski)基因组微卫星资源。基因组DNA经Sau3AⅠ限制性内切酶消化后,选取400―900bp的片段进行PCR全基因组扩增,并利用生物素标记的(CA)12、(GA)12探针进行微卫星片段的富集。将得到的片段与pGEM-T载体连接后转入DH5α大肠杆菌中,然后利用γ-32P标记的放射性同位素探针进行第二次杂交。结果,共获得微卫星基因组文库1600个菌,杂交前菌落PCR检测阳性克隆率为90.91%;杂交后得到的阳性克隆为1300个,占87.25%。从中挑出196个进行测序,192(97.96%)个含有微卫星序列。在得到的微卫星序列中,重复单元除CA/GT、GA/CT外,还观察到单碱基、四碱基、五碱基重复单元。根据侧翼序列应用引物设计软件PrimerPremier5.0设计引物70对,选择合成32对,通过优化PCR反应条件,结果有28对引物可扩增出清晰可重复的目的条带。本研究旨在对黑斑狗鱼基因组资源的开发利用起到一定的促进作用,并为黑斑狗鱼养殖品系的优化、遗传多样性的检测及遗传图谱的构建等奠定基础。     

A protocol for the construction of microsatellite enriched genomic library

An improved protocol for constructing microsatellite-enriched libraries was developed. The procedure depends on digesting genomic DNA with a restriction enzyme that generates blunt-ends, and on ligating linkers that, when dimerized, create a restriction site for a different blunt-end producing restriction enzyme. Efficient ligation of linkers to the genomic DNA fragments is achieved by including restriction enzymes in the ligation reaction that eliminate unwanted ligation products. After ligation, the reaction mixture is subjected to subtractive hybridization without purification. DNA fragments containing microsatellites are captured by biotin-labeled oligonucleotide repeats and recovered using streptavidin-coated beads. The recovered fragments are amplified by PCR using the linker sequence as primer, and cloned directly into a plasmid vector. The linker has the sequence GTTT on the 5′ end, which promotes efficient adenylation of the 3′ ends of the PCR products. Consequently, the amplified fragments could be cloned into vectors without purification. This procedure enables efficient enrichment and cloning of microsatellite sequences, resulting in a library with a low level of redundancy.     

Undermethylated DNA as a source of microsatellites from a conifer genome.

Developing microsatellites from the large, highly duplicated conifer genome requires special tools. To improve the efficiency of developing Pinus taeda L. microsatellites, undermethylated (UM) DNA fragments were used to construct a microsatellite-enriched copy library. A methylation-sensitive restriction enzyme, McrBC, was used to enrich for UM DNA before library construction. Digested DNA fragments larger than 9 kb were then excised and digested with RsaI and used to construct nine dinucleotide and trinucleotide libraries. A total of 1016 microsatellite-positive clones were detected among 11 904 clones and 620 of these were unique. Of 245 primer sets that produced a PCR product, 113 could be developed as UM microsatellite markers and 70 were polymorphic. Inheritance and marker informativeness were tested for a random sample of 36 polymorphic markers using a three-generation outbred pedigree. Thirty-one microsatellites (86%) had single-locus inheritance despite the highly duplicated nature of the P. taeda genome. Nineteen UM microsatellites had highly informative intercross mating type configurations. Allele number and frequency were estimated for eleven UM microsatellites using a population survey. Allele numbers for these UM microsatellites ranged from 3 to 12 with an average of 5.7 alleles/locus. Frequencies for the 63 alleles were mostly in the low-common range; only 14 of the 63 were in the rare allele (q < 0.05) class. Enriching for UM DNA was an efficient method for developing polymorphic microsatellites from a large plant genome.     

Sequence-tagged microsatellite profiling (STMP): a rapid technique for developing SSR markers

We describe a technique, sequence-tagged microsatellite profiling (STMP), to rapidly generate large numbers of simple sequence repeat (SSR) markers from genomic or cDNA. This technique eliminates the need for library screening to identify SSR-containing clones and provides an ~25-fold increase in sequencing throughput compared to traditional methods. STMP generates short but characteristic nucleotide sequence tags for fragments that are present within a pool of SSR amplicons. These tags are then ligated together to form concatemers for cloning and sequencing. The analysis of thousands of tags gives rise to a representational profile of the abundance and frequency of SSRs within the DNA pool, from which low copy sequences can be identified. As each tag contains sufficient nucleotide sequence for primer design, their conversion into PCR primers allows the amplification of corresponding full-length fragments from the pool of SSR amplicons. These fragments permit the full characterisation of a SSR locus and provide flanking sequence for the development of a microsatellite marker. Alternatively, sequence tag primers can be used to directly amplify corresponding SSR loci from genomic DNA, thereby reducing the cost of developing a microsatellite marker to the synthesis of just one sequence-specific primer. We demonstrate the utility of STMP by the development of SSR markers in bread wheat.     

Sequence-tagged microsatellite profiling (STMP): a rapid technique for developing SSR markers

We describe a technique, sequence-tagged microsatellite profiling (STMP), to rapidly generate large numbers of simple sequence repeat (SSR) markers from genomic or cDNA. This technique eliminates the need for library screening to identify SSR-containing clones and provides an approximately 25-fold increase in sequencing throughput compared to traditional methods. STMP generates short but characteristic nucleotide sequence tags for fragments that are present within a pool of SSR amplicons. These tags are then ligated together to form concatemers for cloning and sequencing. The analysis of thousands of tags gives rise to a representational profile of the abundance and frequency of SSRs within the DNA pool, from which low copy sequences can be identified. As each tag contains sufficient nucleotide sequence for primer design, their conversion into PCR primers allows the amplification of corresponding full-length fragments from the pool of SSR amplicons. These fragments permit the full characterisation of a SSR locus and provide flanking sequence for the development of a microsatellite marker. Alternatively, sequence tag primers can be used to directly amplify corresponding SSR loci from genomic DNA, thereby reducing the cost of developing a microsatellite marker to the synthesis of just one sequence-specific primer. We demonstrate the utility of STMP by the development of SSR markers in bread wheat.     

背角无齿蚌基因组(GT)n微卫星DNA特征

利用磁珠富集法筛选背角无齿蚌(Anodonta woodiana)的微卫星分子标记,采用Sau3A1酶对完整DNA进行酶切,以生物素标记的(GT)15寡核苷酸探针从酶切片段中筛选微卫星序列.洗脱的杂交片段克隆到PGEM-T载体上构建富集微卫星基因组文库后,通过菌液PCR筛选检测出阳性克隆进行测序.结果表明:在筛选的18...     

Isolation and enrichment of human genomic CpG islands by methylation-sensitive mirror orientation selection

CpG islands (CGIs) in human genomic DNA are GC-rich fragments whose aberrant methylation is associated with human disease development. In the current study, methylation-sensitive mirror orientation selection (MS-MOS) was developed to efficiently isolate and enrich unmethylated CGIs from human genomic DNA. The unmethylated CGIs prepared by the MS-MOS procedure subsequently were used to construct a CGI library. Then the sequence characteristics of cloned inserts of the library were analyzed by bioinformatics tools, and the methylation status of CGI clones was analyzed by HpaII PCR. The results showed that the MS-MOS method could be used to isolate up to 0.001% of differentially existed unmethylated DNA fragments in two complex genomic DNA. In the CGI library, 34.1% of clones had insert sequences satisfying the minimal criteria for CGIs. Excluding duplicates, 22.0% of the 80,000 clones were unique CGI clones, representing 60% of all the predicted CGIs (about 29,000) in human genomic DNA, and most or all of the CGI clones were unmethylated in human normal cell DNA based on the HpaII PCR analysis results of randomly selected CGI clones. In conclusion, MS-MOS was an efficient way to isolate and enrich human genomic CGIs. The method has powerful potential application in the comprehensive identification of aberrantly methylated CGIs associated with human tumorigenesis to improve understanding of the epigenetic mechanisms involved.     

Highly Informative Single-Copy Nuclear Microsatellite DNA Markers Developed Using an AFLP-SSR Approach in Black Spruce (Picea mariana) and Red Spruce (P. rubens)

Background

Microsatellites or simple sequence repeats (SSRs) are highly informative molecular markers for various biological studies in plants. In spruce (Picea) and other conifers, the development of single-copy polymorphic genomic microsatellite markers is quite difficult, owing primarily to the large genome size and predominance of repetitive DNA sequences throughout the genome. We have developed highly informative single-locus genomic microsatellite markers in black spruce (Picea mariana) and red spruce (Picea rubens) using a simple but efficient method based on a combination of AFLP and microsatellite technologies.

Principal Findings

A microsatellite-enriched library was constructed from genomic AFLP DNA fragments of black spruce. Sequencing of the 108 putative SSR-containing clones provided 94 unique sequences with microsatellites. Twenty-two of the designed 34 primer pairs yielded scorable amplicons, with single-locus patterns. Fourteen of these microsatellite markers were characterized in 30 black spruce and 30 red spruce individuals drawn from many populations. The number of alleles at a polymorphic locus ranged from 2 to 18, with a mean of 9.3 in black spruce, and from 3 to 15, with a mean of 6.2 alleles in red spruce. The polymorphic information content or expected heterozygosity ranged from 0.340 to 0.909 (mean = 0.67) in black spruce and from 0.161 to 0.851 (mean = 0.62) in red spruce. Ten SSR markers showing inter-parental polymorphism inherited in a single-locus Mendelian mode, with two cases of distorted segregation. Primer pairs for almost all polymorphic SSR loci resolved microsatellites of comparable size in Picea glauca, P. engelmannii, P. sitchensis, and P. abies.

Significance

The AFLP-based microsatellite-enriched library appears to be a rapid, cost-effective approach for isolating and developing single-locus informative genomic microsatellite markers in black spruce. The markers developed should be useful in black spruce, red spruce and other Picea species for various genetics, genomics, breeding, forensics, conservation studies and applications.