扎如寺

宝镜岩下,翡翠河旁,绿荫掩映中有一片飞阁流丹,金光耀眼,肃穆巍峨的建筑群,这就是九寨沟内唯一的宗教活动场所——扎如寺。扎如寺建于明末,历史悠久。原占地面积为1.5万平方米,结构精巧,特点突出,具民族风格。整个建筑群包括大殿、藏经楼、乐台、茶房、库房、迎客     

用正交试验法优选二化螟人工饲料配方

采用15因子2水平正交试验,优选出1种二化螟Chilo suppressalis(Walker)人工饲料配方,筛选出影响二化螟发育的5个关键因子,分别是稻茎粉、麦芽粉、稻糠粉、酵母粉和蔗糖。其中稻茎粉、麦芽粉、酵母粉和稻糠粉影响二化螟的幼虫成活率,蔗糖和麦芽粉影响二化螟的幼虫历期,稻茎粉影响二化螟的蛹重。     

粉拟青霉种内nrDNA ITS分析

通过对20株粉拟青霉(Paecilomyces farinosus)ITS1-5．8S-ITS2(rDNA)区域序列测定,确定了粉拟青霉ITS序列,而韩国学者测定的粉拟青霉ITS序列应为细脚拟青霉(P．tenuipes)的序列。序列比较发现,韩国2株未定名的拟青霉(Paecilomyces spp．)菌株(KACC40219、KACC40221)应为粉拟青霉。基于本研究构建的邻接树推断,粉拟青霉的有性型可能是一种虫草。粉拟青霉的起源应为单源的。不同的粉拟青霉菌株的ITS序列具有多态性,源于同一地区的菌株的ITS变异也较大。ITS序列的证据表明,粉拟青霉菌株间的差异与地理来源及寄主均无相关性。     

饲养瓢虫的替代食物:利用地中海粉螟卵与油菜花粉连续多代饲喂稻红瓢虫

稻红瓢虫Micraspis discolor(Fabricius)是稻田害虫捕食性天敌优势种之一,捕食包括蓟马、稻蚜、叶蝉、飞虱Delphacidae、鳞翅目幼虫和卵等多种水稻害虫,对褐飞虱Nilaparvata lugens有控制作用.为了筛选出稻红瓢虫系统化饲养适宜的替代食物,本研究利用地中海粉螟Ephestia kuehniella Zeller卵和油菜花粉分别连续4代饲养稻红瓢虫,分析各代不同食物处理后幼虫各龄的历期、死亡率、雌性比、雌雄体重间的差异,同时也对比了两种处理F2代间雌虫的繁殖力及雌雄成虫的寿命.实验发现,食用地中海粉螟卵的稻红瓢虫比食用油菜花粉的表现力更佳,具体表现为稻红瓢虫取食地中海粉螟卵的幼虫各代历期显著短于取食油菜花粉的处理(P<0.05),F1～F3代取食地中海粉螟卵的稻红瓢虫比取食油菜花粉的稻红瓢虫具有较高的雌性比例(38.46％ ～41.07％vs 50.75％ ～55.56％),取食地中海粉螟卵的各代初羽化稻红瓢虫体重始终大于取食油菜花粉的各代稻红瓢虫.油菜花粉饲喂的F2代雌性稻红瓢虫产卵前期大约是同期地中海粉螟卵处理的2倍多(P<0.05),且稻红瓢虫取食地中海粉螟卵后的最初30天产卵量大于油菜花粉处理,喂饲地中海粉螟卵的F2代稻红瓢虫雌雄成虫寿命比喂饲油菜花粉的F2代稻红瓢虫长.综合分析幼虫发育和繁殖力结果,地中海粉螟卵比油菜花粉更适合稻红瓢虫的生长发育,稻红瓢虫在连续多代饲喂地中海粉螟卵后比饲喂油菜花粉有着更稳定的幼虫发育指标,表明地中海粉螟卵比油菜花粉更适合作为稻红瓢虫系统化扩繁的替代食物.     

宁夏粉蚧科昆虫研究（半翅目：蚧总科）

记述宁夏同族自治区粉蚧科Pseudococcidae昆虫24属41种,其中包括2新种:宁夏草粉蚧Fonscolombia ningxiana,sp.nov.和冰草长粉蚧Longicoccus agropyri.sp.nov.;2中国新记录种:多管刺粉蚧spinococcus multitubulatus(Danzig,1980)和孤独条粉蚧Trionymus singularis Schmutterer,1952:1新组合:赖草长粉蚧Longicoccus leymicola(Tang,1992),comb.n.(移自少粉蚧属Mirococcus Borchsenius);Puto jarudensis Tang(1992)为Ceroputopilosellae Sulc.1898的新异名.模式标本保存在山西农业大学蚧虫研究中心.     

粉垄耕作对耕地土壤酶活性、微生物群落结构和功能多样性的影响

粉垄耕作是中国的一种新型耕作技术,对耕地质量和作物增产有重要影响。设置传统耕作深度20 cm(CK)、粉垄耕作深度35 cm(FL1)和粉垄耕作深度50 cm(FL2)对玉米耕地进行处理,重点研究了粉垄耕作技术对土壤微生态的影响,并阐明土壤微生物群落组成及功能对粉垄耕作的响应。结果表明,FL1、FL2和CK处理玉米产量分别为8.58、8.38和6.22 t/hm~2,FL1和FL2处理增产率分别为34.7%—37.9%。在0—20、20—40 cm土层中,粉垄耕作两个处理的土壤酶活性、微生物群落多样性和功能多样性均显著高于CK处理。通过结构方程模型发现,粉垄耕作直接提高了土壤酶活性、细菌参与养分循环的功能基团和细菌的群落结构,并通过细菌群落间接影响了真菌群落,增加了真菌参与养分循环的功能基团和真菌群落多样性,使土壤微生物碳源利用的能力和功能多样性指数得到提升,以FL1效果更佳。总之,研究从微生物的角度解释了粉垄耕作对土壤微生态的影响机制,为粉垄耕作提升土壤耕地质量提供了理论依据。     

风味冲调紫甘薯粉配方研究

以紫甘薯全粉为主料,以燕麦粉、小米粉、果粉和糖粉为辅料,采用U10（1010）均匀设计,研究了风味冲调紫甘薯粉不同配方的感官性状。结果显示：对感官性状影响最显著的因子是小米粉,其次是燕麦粉、糖粉、柠檬粉,鲜橙粉对感官性状影响很小,几乎可以忽略不计。经过对感官性状的回归分析,结果表明：风味冲调紫甘薯粉最适宜的配方为：紫甘薯粉64~68g、小米粉8~10g、燕麦粉7~9g、柠檬粉6~8g、鲜橙粉1~2g、糖粉9~10g。该配方兼顾了青少年和中老年的口味,具有较好的普适性。     

赣南脐橙粉虱类害虫种类及种群动态研究

通过在赣州开展为期2年的系统调查,明确了为害赣南脐橙的粉虱种类和发生动态。经鉴定,为害赣南脐橙的粉虱为柑橘粉虱Dialeurodes citri Ashmead和黑刺粉虱Aleurocanthus spiniferus Quaintance。2013年柑橘粉虱,黑刺粉虱卵、若虫、伪蛹、成虫数量平均密度分别为(7.61,6.67,5.45,47.43)头/叶(板),(3.84,2.84,1.82,31.80)头/叶(板);2014年柑橘粉虱,黑刺粉虱分别为(8.29,6.92,5.18,27.16)头/叶(板),(7.89,6.18,4.35,24.06)头/叶(板)。2013-2014年,两种粉虱发生数量较大,全年有多个发生高峰。柑橘粉虱和黑刺粉虱是赣南脐橙上的重要害虫,需加强防治工作,控制为害。     

重金属铅对椭圆食粉螨形态及SOD、GST活性的影响

【目的】Pb污染严重影响生物生长发育和繁殖,本文研究12.5、25、50、100mg/kg4种Pb2+浓度下椭圆食粉螨形态特征及SOD、GST活性的变化规律,探究粉螨对重金属胁迫的响应机制。【方法】以不加Pb为对照,测量不同浓度Pb2+胁迫下,不同螨态椭圆食粉螨形态特征长度及SOD、GST酶活。【结果】Pb胁迫对椭圆食粉螨体型大小整体表现为抑制作用,但随Pb2+浓度升高呈现先增大后减小的趋势。抗氧化酶SOD、GST酶活性呈现先升高后下降,整体仍对椭圆食粉螨SOD、GST酶活性产生促进作用。【结论】Pb2+胁迫对椭圆食粉螨体型大小整体表现为抑制作用,SOD、GST活性产生促进作用。椭圆食粉螨SOD、GST活性可用于监测储藏作物重金属污染。     

基于MaxEnt模型的小巢粉虱在中国的潜在地理分布

小巢粉虱是一种危险性外来入侵害虫,现已传入我国香港、海南。阐明小巢粉虱在我国的潜在地理分布及其主要限制环境因子,可为该虫早期预警与防控提供理论依据。本文利用MaxEnt模型对小巢粉虱在我国的潜在适生区进行预测,使用刀切法及环境变量响应曲线对影响小巢粉虱分布的环境因子进行评估。结果表明:小巢粉虱在我国的潜在适生区分布于海南、台湾、广东、广西、福建、云南、贵州、湖南、江西、浙江、上海、安徽、湖北、重庆、四川、西藏等省区;适生区总面积224.9万km~2,约占我国国土面积的23.4%,其中高度适生区面积38.2万km~2,中度适生区面积47.6万km~2,低度适生区面积139.1万km~2;最冷月最低温(bio06)、最干季平均气温(bio09)、最冷季降水量(bio19)是影响小巢粉虱潜在地理分布的主导环境因子。我国各潜在适生区、特别是华南沿海地区应加强检疫,防止小巢粉虱进一步扩散。     

Construction and characterization of a normalized yeast two-hybrid library derived from a human protein-coding clone collection

The nuclear yeast two-hybrid (Y2H) system is the most widely used technology for detecting interactions between proteins. A common approach is to screen specific test proteins (baits) against large compilations of randomly cloned proteins (prey libraries). For eukaryotic organisms, libraries have traditionally been generated using messenger RNA (mRNA) extracted from various tissues and cells. Here we present a library construction strategy made possible by ongoing public efforts to establish collections of full-length protein encoding clones. Our approach generates libraries that are essentially normalized and contain both randomly fragmented as well as full-length inserts. We refer to this type of protein-coding clone-derived library as random and full-length (RAFL) Y2H library. The library described here is based on clones from the Mammalian Gene Collection, but our strategy is compatible with the use of any protein-coding clone collection from any organism in any vector and does not require inserts to be devoid of untranslated regions. We tested our prototype human RAFL library against a set of baits that had previously been searched against multiple cDNA libraries. These Y2H searches yielded a combination of novel as well as expected interactions, indicating that the RAFL library constitutes a valuable complement to Y2H cDNA libraries.     

A surface display yeast two-hybrid screening system for high-throughput protein interactome mapping

Despite the wide acceptance of yeast two-hybrid (Y2H) system for protein-protein interaction analysis and discovery, conventional Y2H assays are not well suited for high-throughput screening of the protein interaction network (“interactome”) on a genomic scale due to several limitations, including labor-intensive agar plating and colony selection methods associated with the use of nutrient selection markers, complicated reporter analysis methods associated with the use of LacZ enzyme reporters, and incompatibility of the liquid handling robots. We recently reported a robust liquid culture Y2H system based on quantitative analysis of yeast-enhanced green fluorescent protein (yEGFP) reporters that greatly increased the analysis throughput and compatibility with liquid handling robots. To further advance its utility in high-throughput complementary DNA (cDNA) library screening, we report the development of a novel surface display Y2H (sdY2H) library screening system with uniquely integrated surface display hemagglutination (sdHA) antigen and yEGFP reporters. By introduction of a surface reporter sdHA into the yEGFP-based Y2H system, positive Y2H targets are quickly isolated from library cells by a simple magnetic separation without a large plating effort. Moreover, the simultaneous scoring of multiple reporters, including sdHA, yEGFP, and conventional nutrient markers, greatly increased the specificity of the Y2H assay. The feasibility of the sdY2H assay on large cDNA library screening was demonstrated by the successful recovery of positive P53/T interaction pairs at a target-to-background ratio of 1:1,000,000. Together with the massive parallel DNA sequencing technology, it may provide a powerful proteomic tool for high-throughput interactome mapping on a genomic scale.     

Construction of a reading frame-independent yeast two-hybrid vector system for site-specific recombinational cloning and protein interaction screening

The yeast two-hybrid (Y2H) system is a powerful method to identify protein-protein inter-actions (PPI) in vivo, requiring minimal prior information of the putative interactors. The time and effort required for each experiment can be significantly reduced if the "bait" and the "prey" proteins are cloned into specific recombination-amenable two-hybrid vectors. We describe the construction of a reading frame-independent vector system for Y2H PPI studies. The described vector system knits together the advantages of site-specific recombination cloning with the Y2H system. The produced plasmids enable recombination-based cloning of genes or gene fragments in all possible reading frames into Y2H library vectors. Thus, Y2H screening libraries can be rapidly constructed and will present more amino termini in the correct reading frame. Additionally, advantageous for small-scale Y2H studies, there is no need to know the natural reading frame of the genes of interest, because the bait and prey genes can be transferred into the vectors by a single reaction and are present in all possible reading frames. Since the Y2H system per se is a positive selection system, only pairs of bait and prey genes harboring the correct reading frames will emerge. We tested the new vectors within the Y2H system and demonstrated full functionality without any undesired effects on the Y2H system itself. Besides the vector construction, we investigated the utility of the system for Y2H analysis and demonstrated clearly its practicability in genome-wide Y2H screenings and the advantage of using additional reading-frame Y2H cDNA libraries. We performed a series of genome-wide Y2H library screenings with the human vitamin D receptor protein (VDR) as bait. We investigated: (i) whether more protein interactors are found by using three instead of one reading-frame destination vectors; (ii) how much overlap between the different reading-frame libraries exists; and (iii) the rate of possible additional autoactivators. We conclude that our vectors deliver significantly more interactors and outperform a single reading-frame library. This new system could enable simple and fast large-scale PPI studies and the construction of high-quality screening libraries.     

A facile route to dynamic glycopeptide libraries based on disulfide-linked sugar-peptide coupling

We report here that disulfide-linked dynamic glycopeptide libraries can be constructed from 1-thiosugar and cysteine-rich oligopeptide building blocks upon gentle air oxidation of a slightly basic (pH 7.8) aqueous solution thereof. A mixture of 1-thiogalactose and two oligopeptides H2N-CysGlyCysGly-CO2H and H2N-GlyCycCysGlyGly-CO2H, for example, affords a poorly HPLC-resolved disulfide library composed of various sugar-peptide conjugates and cyclic peptides, at least 10 of which can be identified by ESI mass spectrometry. The building components of disulfide members are exchangeable with each other in the presence of dithiothreitol as an initiator to allow dynamic equilibration. A preliminary SPR examination shows that the thiogalactose-derived library indeed contains active divalent galactoside species capable of cross-linking peanut lectin molecules.     

Design and Selection of a Camelid Single-Chain Antibody Yeast Two-Hybrid Library Produced De Novo for the Cap Protein of Porcine Circovirus Type 2 (PCV2)

Nanobodies (or variable domain of the heavy chain of the heavy-chain antibodies, VHHs) are single-domain antigen-binding fragments derived from camelid heavy chain antibodies. Their comparatively small size, monomeric behavior, high stability, high solubility, and ability to bind epitopes inaccessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. In this paper, for the first time, we created the immunized Camelus Bactrianus VHH yeast two-hybrid (Y2H) library according to the Clontech Mate & Plate library construction system. The transformation efficiency and titer of the VHH Y2H library were 7.26×10⁶ cfu/3 µg and 2×10⁹ cfu/ml, which met the demand for Y2H library screening. Using as an example the porcine circovirus type 2 (PCV2) Cap protein as bait, we screened 21 positive Cap-specific VHH sequences. Among these sequences, 7 of 9 randomly selected clones were strongly positive as indicated by enzyme-linked immunosorbent assay, either using PCV2 viral lysis or purified Cap protein as coated antigen. Additionally, the immunocytochemistry results further indicated that the screened VHHs could specifically detected PCV2 in the infected cells. All this suggests the feasibility of in vivo VHH throughput screening based on Y2H strategy.     

应用噬菌体展示随机肽库淘筛mAb 5H5识别的抗原表位

本文利用噬菌体随机9肽库探索汉滩病毒（HTNV）核衣壳蛋白（NP）B细胞抗原表位。以抗HTNV NP单克隆抗体(mAb）5H5作为筛选分子,生物淘洗噬菌体递呈的随机9肽库。阳性克隆经夹心ELISA、竞争ELISA鉴定后,随机挑取10个克隆,DNA测序,与HTNV76－118株S基因进行同源性分析。结果显示筛选到的噬菌体能特异地与5H5结合,这种结合可被天然抗原所抑制。10个克隆的氨基酸序列相同,均为VRDAEEQYE,与76－118株NP氨基端的aa25-33一致。证实了该线性表位是mAb 5H5识别的表位,噬菌体肽库有助于病毒抗原表位的确定。     

Protective effect of vaccination with DNA of the H. Pylori genomic library in experimentally infected mice

Immunologically mediated protection against H. pylori infection is an attractive alternative to antibiotic treatment. We compared the efficacy of conventional protein vaccination with that of genetic vaccination against experimental infection with H. pylori in mice. For oral immunization, we used the recombinant peptide of an antigenic fragment of UreB (rUreB) or H. pylori-whole cell lysate antigens, and for genetic immunization, we used recombinant pcDNA and pSec plasmids inserted with the fragment of ureB or DNA of the H. pylori genomic library. Mice were challenged with the mouse stomach-adapted H. pylori Sidney Strain. The detection of gastric bacterial colonization was performed by real-time PCR of a 26-kDa Helicobacter-specific gene, and the presence of serum H. pylori-specific antibodies was determined using direct ELISA assay. The most effective treatment appeared to be oral vaccination with rUreB and either intramuscular or intradermal vaccination with DNA of the H. pylori genomic library. Intradermal genetic vaccination with genomic library DNA significantly increased the IgG antibody response. Our study revealed acceptable efficacies of genetic vaccination with DNA of the H. pylori genomic library.     

Isolation and characterization of two human H1 histone genes within clusters of core histone genes

Two human H1 histone genes, termed H1.3 and H1.4, were isolated from two cosmid clones. The H1.4 gene is associated with an H2B gene, whereas genes coding for all four core histones are located in the vicinity of the H1.3 gene. This cluster arrangement was found both in the two cosmid clones and on overlapping bacteriophage clones isolated from an EMBL3 library. In continuation of our previous analysis of two human H1 genes, this analysis raises the number of completely sequenced H1 histone genes within clusters of core histone genes to four.     

Repeated Random Mutagenesis of alpha-Amylase from Bacillus licheniformis for Improved pH Performance

The alpha-amylases activity was improved by random mutagenesis and screening. A region comprising residues from the position 34-281 was randomly mutated in B. licheniformis alpha-amylase (AmyL), and the library with mutations ranging from low, medium, and high frequencies was generated. The library was screened using an effective liquid-phase screening method to isolate mutants with an altered pH profile. The sequencing of improved variants indicated 2-5 amino acid changes. Among them, mutant TP8H5 showed an altered pH profile as compared with that of wild type. The sequencing of variant TP8H5 indicated 2 amino acid changes, Ile157Ser and Trp193Arg, which were located in the solvent accessible flexible loop region in domain B.     

Engineering antibody heavy chain CDR3 to create a phage display Fab library rich in antibodies that bind charged carbohydrates

A number of small charged carbohydrate moieties have been associated with inflammation and cancer. However, the development of therapeutic Abs targeting these moieties has been hampered by their low immunogenicity and their structural relationship to self-Ag. We report the design of an Ab repertoire enriched in Abs binding to small charged carbohydrates and the construction of a human Fab phagemid library, "FAB-CCHO." This library combines L chain Ig sequences from human donors and H chain synthetic diversity constructed in key Ag contact sites in CDRs 1, 2, and 3 of the human framework V(H)3-23. The H chain CDR3 has been engineered to enrich the library in Abs that bind charged carbohydrates by the introduction of basic residues at specific amino acid locations. These residues were selected on the basis of anti-carbohydrate Ab sequence alignment. The success of this design is demonstrated by the isolation of phage Abs against charged carbohydrate therapeutic target Ags such as sulfated sialyl-Lewis X glycan and heparan sulfate.