凡纳滨对虾分子标记筛选、连锁图谱构建和QTL定位

利用AFLP和微卫星标记，以凡纳滨对虾F1全同胞家系为作图群体，构建了凡纳滨对虾的雌性和雄性遗传连锁图谱并对生长相关性状体长和体重进行了QTL分析。利用经过筛选的108对AFLP引物组合，对亲本和94个子代个体进行了分离分析。共得到2041个多态AFLP标记（1:1分离）。平均每个引物组合产生20个多态片段。有826个AFLP标记偏离孟德尔遗传（40.5%, P<0.05）。所筛选的100个微卫星标记中有30个在家系中有作图信息，分别有24个和20个位点可以用于母本和父本的作图。对父母本β-1，3-葡聚糖结合蛋白(BGBP)，脂多糖和葡聚糖结合蛋白(LGBP)和蜕皮抑制激素（MIH）等基因片段进行序列分析得到一些可用于构建连锁图谱的SNP标记。 对所有的分离标记进行了连锁分析，分别绘制了凡纳滨对虾的雌性和雄性连锁图谱。雌性连锁图谱的框架图有319个遗传标记组成，分布于45个连锁群，其中AFLP标记300个，微卫星位点18个，性别标记一个，连锁群长度从29.5cM到260.0cM, 每个连锁群含4－16个标记。图谱长度为4134.4 cM, 各连锁群平均图距在7.6到25.9 cM之间，总平均图距为15.1 cM。有267个标记（含14个微卫星）整合到雄性框架图上。 雄性框架图也含45个连锁群，长度从14.2cM到161.1cM, 图谱长度为3220.9 cM, 各连锁群平均图距在4.1到25.5 cM之间，总平均图距为14.5 cM。 作图群体所有个体的性别均作为标记整合到雌雄分离信息中，在94个F1个体中，54个为雌虾，40个是雄虾。在雌性图谱第29连连锁群上，性别与三个微卫星标记紧密连锁（v1f148, v145f120, v95f166）, 图距分别为6.6, 8.6 和8.6cM, 相应LOD值分别为17.8 14.3 和 16.4。 与雌性图相对应的是，雄性图谱未发现任何与性别连锁的标记。推测凡纳滨对虾的性别决定机制可能是ZW型，其中雌性为异配性别。 体长和体重均显示出连续变异的特点，显示这些与生长相关的性状都是典型的数量性状或多基因遗传。体重和体长符合正态分布且两性状之间存在着显著的相关性（P<0.001），Pearson 相关系数为0.95。在凡纳滨对虾雌性图谱和雄性图谱上，共定位了6个与体长和体重等生长性状相关的QTL，可解释的表型变异从15.1％到21.3％。共检测到3个与体长相关的QTL，包括两个正向效应的QTL和一个负向的QTL，分别定位到雌性的第6连锁群和雄性的第4和11 连锁群上。体重也检测到3个QTL，其中一个QTL加性效应是正向的，分别位于雌性的第6和7连锁群及雄性的第11连锁群上。QTL主要集中在雌性图谱的Fc6和Fc7及雄性图的Mc4和Mc11上。体长和体重的QTL定位结果中各有两个是十分相似的，其相邻的标记位点完全一样，只是相应的LOD值略有差异，还各存在两性状特异的一个QTL。大多QTL与相邻标记之间的距离只有0.1cM(仅Wfc7的距离较大)，为进一步的精细定位奠定了基础。 分子标记筛选、遗传图谱的构建及生长相关性状的QTL定位为我们下一步从事分子标记辅助选择育种,QTL精细定位和比较基因作图打下基础，并最终推动凡纳滨对虾的遗传改良。

肉食性鱼类补偿生长特性的研究

本文主要以花鲈[Lateolabrax．japonicus (C．& V．)]和褐牙鲆[Paralichthys olivaceus(T. & S.)]作为海洋肉食性鱼类的代表种类，根据鱼类生态生理学理论，通过设定不同饥饿时问下因子水平，研究两种鱼类摄食率、排粪率、转化效率和SGR等生态生理效率的状态变化。其目的在于研究高营养级鱼类在海洋生态系统中的下行控制作用(Top-down Effect)，以及肉食性鱼类的生态对策与鱼类资源补充机制的相互关系，为深入解析鱼类资源生产力及其持续利用海洋生物资源，提供科学依据。其主要研究结果概述如下：1．花鲈： 饥饿0(对照组)、4、8、12、16d后恢复投喂，过量投喂淡水桡足类，温度为19.5±2.0℃，初始体重为0.61-0.93g，平均体重为0．79g实验周期为28d。1.1饥饿时间对花鲈的体重损失率产生显著影响，受过饥饿的个体的湿重失率 (LR_W)与饥饿时间(t)的关系为：LR_W=2.2164t-3.6634(r~2=0.9767，p13d)的个体的排粪率显著大于其它各组，其它各组间排粪率差异不显著；2.2在整个实验周期36d内，对照组与经受3—8d饥饿的实验组的SGR显著大于经受饥饿时间大于lOd的实验组，饥饿3-8d的完全补偿生长，饥饿10-15d 的个体发生部分补偿生长，饥饿18d的个体不能补偿生长；经受饥饿个体的摄食率低于对照组；除了饥饿18d的个体的饵料转化率显著小于对照组外，其它各组的转化率没有显著差异；经受长时间饥饿(>13d)的个体的排粪率显著大于其它各组，其它各组问没有显著的差异。2．3平均体重为2．59g的褐牙鲆，饥饿至不可恢复点(PNR)的时间大于18d。3．花鲈和褐牙鲆在经受饥饿后的体重损失率与饥饿的时间正相关。在实验期间，花鲈体重损失率与饥饿时间成一次线性关系，褐牙鲆体重损失率与饥饿时间成对数函数关系。在饥饿后均会发生补偿生长。3.1比较研究结果，表明在适当时间饥饿后的个体均会发生补偿生长，其超生长速度与饥饿时间密切相关。花鲈在饥饿至不可恢复点的临界状态时发生最强烈的超生长现象，而褐牙鲆则是在饥饿3—5d后恢复投喂时可以观测到最高速度的超生长。3.2经受适当时间饥饿后的个体在短期内，其摄食率大于对照组的摄食强度。曾经饥饿的时间越长，花鲈在饥饿结束后的摄食率越高，褐牙鲆在饥饿结束后的高摄食率持续的时间越长。3.3经受适当时间饥饿后的个体，在短期内的转化率均会大于对照组的转化 效率，且随饥饿时间而加强；高转化效率持续的时间与饥饿时间无关。褐牙鲆的高转化率持续的时间很短，且很快下降至低于对照组的水平，其下降程度与饥饿时问成正相关，经受过度饥饿的花鲈和褐牙鲆转化率显著低于对照组。3．4花鲈的补偿生长是饵料转化率提高的结果；而褐牙鲆的补偿生长的原因则因其经受的饥饿程度而异。从饥饿后恢复投喂较短的时间周期(4d和6d)看，花鲈和褐牙鲆的补偿生长是转化率和摄食率共同提高的结果；但从饥饿后恢复投喂较长的时问周期(12d和1 8d)看，经受短期(4-8d)饥饿的花鲈的补偿生长是转化率显著提高的结果，而经受长期(12d)饥饿的花鲈的补偿生长是转化率显著提高的结果，褐牙鲆则是摄食率显著提高的结果。3．5从整个实验周期来看，对花鲈的饥饿降低了其生长速度和饵料转化率，但对褐牙鲆的短期饥饿(<8d)不会降低其生长速度，也不会影响一定时间(3-13d)饥饿的个体的饵料转化率。

Identification of quantitative trait loci for growth-related traits in the Pacific abalone Haliotis discus hannai Ino

The locations and effects of quantitative trait loci (QTL) were estimated for nine characters for growth-related traits in the Pacific abalone (Haliotis discus hannai Ino) using a randomly amplified polymorphic DNA (RAPD), amplification fragment length polymorphism (AFLP) and SSR genetic linkage map. Twenty-eight putatively significant QTLs (LOD > 2.4) were detected for nine traits (shell length, shell width, total weight, shell weight, weight of soft part, muscle weight, gonad and digestive gland weight, mantle weight and gill weight). The percentage of phenotypic variation explained by a single QTL ranged from 8.0% to 35.9%. The significant correlations (P < 0.001) were found among all the growth-related traits, and Pearson's correlation coefficients were more than 0.81. For the female map, the QTL for growth were concentrated on groups 1 and 4 linkage maps. On the male map, the QTL that influenced growth-related traits gathered on the groups 1 and 9 linkage maps. Genetic linkage map construction and QTL analysis for growth-related traits are the basis for the marker-assisted selection and will eventually improve production and quality of the Pacific abalone.

Genetic mapping of size-related quantitative trait loci (QTL) in the bay scallop (Argopecten irradians) using AFLP and microsatellite markers

We constructed genetic linkage maps for the bay scallop Argopecten irradians using AFLP and microsatellite markers and conducted composite interval mapping (CIM) of body-size-related traits. Three hundred seventeen AFLP and 10 microsatellite markers were used for map construction. The female parent map contained 120 markers in 15 linkage groups, spanning 479.6 cM with an average interval of 7.0 cM. The male parent map had 190 markers in 17 linkage groups, covering 883.8 cM at 7.2 cM per marker. The observed coverage was 70.4% for the female and 81.1% for the male map. Markers that were distorted toward the same direction were closely linked to each other on the genetic maps, suggesting the presence of genes important for survival. Six size-related traits, shell length, shell height, shell width, total weight, soft tissue weight, and shell weight, were measured for QTL mapping. The size data were significantly correlated with each other. We subjected the data, log transformed firstly, to a principle component analysis and use the first principle component for QTL mapping. CIM analysis revealed one significant QTL (LOD=2.69, 1000 permutation, P<0.05) in linkage group 3 on the female parent map. The mapping of size-related QTL in this study raises the possibility of improving the growth of bay scallops through marker-assisted selection. (c) 2007 Published by Elsevier B.V.

Determination of five pharmacologically active compounds extracted from Rhodiola for natural product drug discovery with HPLC-APCI-MS

A rapid and sensitive liquid chromatography-atmospheric pressure chemical ionization mass spectrometry (HPLC-APCI-MS) assay for the determination of five pharmacologically active compounds (PAC) extracted from the traditional Chinese medicine, Rhodiola , namely salidroside, tyrosol, rhodionin, gallic acid, and ethyl gallate has been developed. In this method, PAC could be baseline separated and detected with DAD at 275 nm. The validation of the method, including sensitivity, linearity, repeatability, and recovery, was examined. The linear calibration curves were acquired with correlation coefficient >0.999 and the limits of detection LOD (at a signal-to-noise ratio=3:1) were between 0.058 and 1.500 mu mol/L. It was found, that the amounts of PAC varied with different species of Rhodiola . The established method is rapid and reproducible for the separation of five natural pharmacologically active compounds from extracts of Rhodiola with satisfactory results.

Genome-wide association study for subclinical atherosclerosis in major arterial territories in the NHLBI's Framingham Heart Study

INTRODUCTION:Subclinical atherosclerosis (SCA) measures in multiple arterial beds are heritable phenotypes that are associated with increased incidence of cardiovascular disease. We conducted a genome-wide association study (GWAS) for SCA measurements in the community-based Framingham Heart Study.METHODS:Over 100,000 single nucleotide polymorphisms (SNPs) were genotyped (Human 100K GeneChip, Affymetrix) in 1345 subjects from 310 families. We calculated sex-specific age-adjusted and multivariable-adjusted residuals in subjects tested for quantitative SCA phenotypes, including ankle-brachial index, coronary artery calcification and abdominal aortic calcification using multi-detector computed tomography, and carotid intimal medial thickness (IMT) using carotid ultrasonography. We evaluated associations of these phenotypes with 70,987 autosomal SNPs with minor allele frequency [greater than or equal to] 0.10, call rate [greater than or equal to] 80%, and Hardy-Weinberg p-value [greater than or equal to] 0.001 in samples ranging from 673 to 984 subjects, using linear regression with generalized estimating equations (GEE) methodology and family-based association testing (FBAT). Variance components LOD scores were also calculated.RESULTS:There was no association result meeting criteria for genome-wide significance, but our methods identified 11 SNPs with p < 10-5 by GEE and five SNPs with p < 10-5 by FBAT for multivariable-adjusted phenotypes. Among the associated variants were SNPs in or near genes that may be considered candidates for further study, such as rs1376877 (GEE p < 0.000001, located in ABI2) for maximum internal carotid artery IMT and rs4814615 (FBAT p = 0.000003, located in PCSK2) for maximum common carotid artery IMT. Modest significant associations were noted with various SCA phenotypes for variants in previously reported atherosclerosis candidate genes, including NOS3 and ESR1. Associations were also noted of a region on chromosome 9p21 with CAC phenotypes that confirm associations with coronary heart disease and CAC in two recently reported genome-wide association studies. In linkage analyses, several regions of genome-wide linkage were noted, confirming previously reported linkage of internal carotid artery IMT on chromosome 12. All GEE, FBAT and linkage results are provided as an open-access results resource at http://www.ncbi.nlm.nih.gov/projects/gap/cgi-bin/study.cgi?id=phs000007.CONCLUSION:The results from this GWAS generate hypotheses regarding several SNPs that may be associated with SCA phenotypes in multiple arterial beds. Given the number of tests conducted, subsequent independent replication in a staged approach is essential to identify genetic variants that may be implicated in atherosclerosis.

Genome-wide association with bone mass and geometry in the Framingham Heart Study

BACKGROUND:Osteoporosis is characterized by low bone mass and compromised bone structure, heritable traits that contribute to fracture risk. There have been no genome-wide association and linkage studies for these traits using high-density genotyping platforms.METHODS:We used the Affymetrix 100K SNP GeneChip marker set in the Framingham Heart Study (FHS) to examine genetic associations with ten primary quantitative traits: bone mineral density (BMD), calcaneal ultrasound, and geometric indices of the hip. To test associations with multivariable-adjusted residual trait values, we used additive generalized estimating equation (GEE) and family-based association tests (FBAT) models within each sex as well as sexes combined. We evaluated 70,987 autosomal SNPs with genotypic call rates [greater than or equal to]80%, HWE p [greater than or equal to] 0.001, and MAF [greater than or equal to]10% in up to 1141 phenotyped individuals (495 men and 646 women, mean age 62.5 yrs). Variance component linkage analysis was performed using 11,200 markers.RESULTS:Heritability estimates for all bone phenotypes were 30-66%. LOD scores [greater than or equal to]3.0 were found on chromosomes 15 (1.5 LOD confidence interval: 51,336,679-58,934,236 bp) and 22 (35,890,398-48,603,847 bp) for femoral shaft section modulus. The ten primary phenotypes had 12 associations with 100K SNPs in GEE models at p < 0.000001 and 2 associations in FBAT models at p < 0.000001. The 25 most significant p-values for GEE and FBAT were all less than 3.5 x 10-6 and 2.5 x 10-5, respectively. Of the 40 top SNPs with the greatest numbers of significantly associated BMD traits (including femoral neck, trochanter, and lumbar spine), one half to two-thirds were in or near genes that have not previously been studied for osteoporosis. Notably, pleiotropic associations between BMD and bone geometric traits were uncommon. Evidence for association (FBAT or GEE p < 0.05) was observed for several SNPs in candidate genes for osteoporosis, such as rs1801133 in MTHFR; rs1884052 and rs3778099 in ESR1; rs4988300 in LRP5; rs2189480 in VDR; rs2075555 in COLIA1; rs10519297 and rs2008691 in CYP19, as well as SNPs in PPARG (rs10510418 and rs2938392) and ANKH (rs2454873 and rs379016). All GEE, FBAT and linkage results are provided as an open-access results resource at http://www.ncbi.nlm.nih.gov/projects/gap/cgi-bin/study.cgi?id=phs000007.CONCLUSION:The FHS 100K SNP project offers an unbiased genome-wide strategy to identify new candidate loci and to replicate previously suggested candidate genes for osteoporosis.

Mapping the gene for autosomal dominant restless legs syndrome in an Irish family

Restless Legs Syndrome (RLS) is a common neurological disorder affecting nearly 15% of the general population. Ironically, RLS can be described as the most common condition one has never heard of. It is usually characterised by uncomfortable, unpleasant sensations in the lower limbs inducing an uncontrollable desire to move the legs. RLS exhibits a circadian pattern with symptoms present predominantly in the evening or at night, thus leading to sleep disruption and daytime somnolence. RLS is generally classified into primary (idiopathic) and secondary (symptomatic) forms. Primary RLS includes sporadic and familial cases of which the age of onset is usually less than 45 years and progresses slowly with a female to male ratio of 2:1. Secondary forms often occur as a complication of another health condition, such as iron deficiency or thyroid dysfunction. The age of onset is usually over 45 years, with an equal male to female ratio and more rapid progression. Ekbom described the familial component of the disorder in 1945 and since then many studies have been published on the familial forms of the disorder. Molecular genetic studies have so far identified ten loci (5q, 12q, 14p, 9p, 20p, 16p, 19p, 4q, 17p). No specific gene within these loci has been identified thus far. Association mapping has highlighted a further five areas of interest. RLS6 has been found to be associated with SNPs in the BTBD9 gene. Four other variants were found within intronic and intergenic regions of MEIS1, MAP2K5/LBXCOR1, PTPRD and NOS1. The pathophysiology of RLS is complex and remains to be fully elucidated. Conditions associated with secondary RLS, such as pregnancy or end-stage renal disease, are characterised by iron deficiency, which suggests that disturbed iron homeostasis plays a role. Dopaminergic dysfunction in subcortical systems also appears to play a central role. An ongoing study within the Department of Pathology (University College Cork) is investigating the genetic characteristics of RLS in Irish families. A three generation RLS pedigree RLS3002 consisting of 11 affected and 7 unaffected living family members was recruited. The family had been examined for four of the known loci (5q, 12q, 14p and 9p) (Abdulrahim 2008). The aim of this study was to continue examining this Irish RLS pedigree for possible linkage to the previously described loci and associated regions. Using informative microsatellite markers linkage was excluded to the loci on 5q, 12q, 14p, 9p, 20p, 16p, 19p, 4q, 17p and also within the regions reported to be associated with RLS. This suggested the presence of a new unidentified locus. A genome-wide scan was performed using two microsatellite marker screening sets (Research Genetics Inc. Mapping set and the Applied Biosystems Linkage mapping set version 2.5). Linkage analysis was conducted under an autosomal dominant model with a penetrance of 95% and an allele frequency of 0.01. A maximum LOD score of 3.59 at θ=0.00 for marker D19S878 indicated significant linkage on chromosome 19p. Haplotype analysis defined a genetic region of 6.57 cM on chromosome 19p13.3, corresponding to 2.5 Mb. There are approximately 100 genes annotated within the critical region. Sequencing of two candidate genes, KLF16 and GAMT, selected on the assumed pathophysiology of RLS, did not identify any sequence variant. This study provides evidence of a novel RLS locus in an Irish pedigree, thus supporting the picture of RLS as a genetically heterogeneous trait.

Rapid separation of biologically important low molecular weight compounds by microchip electrophoresis and ultra-fast performance liquid chromatography

The research work in this thesis reports rapid separation of biologically important low molecular weight compounds by microchip electrophoresis and ultrahigh liquid chromatography. Chapter 1 introduces the theory and principles behind capillary electrophoresis separation. An overview of the history, different modes and detection techniques coupled to CE is provided. The advantages of microchip electrophoresis are highlighted. Some aspects of metal complex analysis by capillary electrophoresis are described. Finally, the theory and different modes of the liquid chromatography technology are presented. Chapter 2 outlines the development of a method for the capillary electrophoresis of (R, S) Naproxen. Variable parameters of the separation were optimized (i.e. buffer concentration and pH, concentration of chiral selector additives, applied voltage and injection condition).The method was validated in terms of linearity, precision, and LOD. The optimized method was then transferred to a microchip electrophoresis system. Two different types of injection i.e. gated and pinched, were investigated. This microchip method represents the fastest reported chiral separation of Naproxen to date. Chapter 3 reports ultra-fast separation of aromatic amino acid by capillary electrophoresis using the short-end technique. Variable parameters of the separation were optimized and validated. The optimized method was then transferred to a microchip electrophoresis system where the separation time was further reduced. Chapter 4 outlines the use of microchip electrophoresis as an efficient tool for analysis of aluminium complexes. A 2.5 cm channel with linear imaging UV detection was used to separate and detect aluminium-dopamine complex and free dopamine. For the first time, a baseline, separation of aluminium dopamine was achieved on a 15 seconds timescale. Chapter 5 investigates a rapid, ultra-sensitive and highly efficient method for quantification of histamine in human psoriatic plaques using microdialysis and ultrahigh performance liquid chromatography with fluorescence detection. The method utilized a sub-two-micron packed C18 stationary phase. A fluorescent reagent, 4-(1-pyrene) butyric acid N-hydroxysuccinimide ester was conjugated to the primary and secondary amino moieties of histamine. The dipyrene-labeled histamine in human urine was also investigated by ultrahigh pressure liquid chromatography using a C18 column with 1.8 μm particle diameter. These methods represent one of the fastest reported separations to date of histamine using fluorescence detection.

Influence of the Graphical Levels of Detail of a Virtual Thrower on the Perception of the Movement

Virtual reality has a number of advantages for analyzing sports interactions such as the standardization of experimental conditions, stereoscopic vision, and complete control of animated humanoid movement. Nevertheless, in order to be useful for sports applications, accurate perception of simulated movement in the virtual sports environment is essential. This perception depends on parameters of the synthetic character such as the number of degrees of freedom of its skeleton or the levels of detail (LOD) of its graphical representation. This study focuses on the influence of this latter parameter on the perception of the movement. In order to evaluate it, this study analyzes the judgments of immersed handball goalkeepers that play against a graphically modified virtual thrower. Five graphical representations of the throwing action were defined: a textured reference level (L0), a nontextured level (L1), a wire-frame level (L2), a moving point light display (MLD) level with a normal-sized ball (L3), and a MLD level where the ball is represented by a point of light (L4). The results show that judgments made by goalkeepers in the L4 condition are significantly less accurate than in all the other conditions (p

Variation in RTN3 and PPIL2 Genes Does not Influence Platelet Membrane β-Secretase Activity or Susceptibility to Alzheimer’s Disease in the Northern Irish Population

beta-site amyloid precursor protein cleaving enzyme (BACE1) is the rate-limiting enzyme for production of beta-amyloid peptides (A beta), which are proposed to drive the pathological changes found in Alzheimer's disease (AD). Reticulon 3 (RTN3) is a negative modulator of BACE1 (beta-secretase) proteolytic activity, while peptidylprolyl isomerase (cyclophilin)-like 2 (PPIL2) positively regulates BACE1 expression. The present study investigated whether there was any association between genetic variation in RTN3 and PPIL2, and either risk for AD, or levels of platelet beta-secretase activity, in a large Northern Irish case-control sample. Four hundred and sixty-nine patients with a diagnosis of probable AD (NINCDS-ADRDA criteria) and 347 control individuals (MMSE > 28/30) were genotyped. SNPs in both genes were selected by downloading genotype data from the International HapMap Project (Phase II) and tags selected using multimarker approach in Haploview, where r (2) > 0.8 and LOD > 3.0. Non-synonymous SNPs of interest were also included. Genotyping was performed by Sequenom iPLEX and TaqMan technologies. Alleles, genotypes and multi-marker haplotypes were tested for association with AD, and platelet beta-secretase activities were measured for a subset of individuals (n = 231). Eight SNPs in RTN3 and 7 in PPIL2 were genotyped. We found no significant associations between allele, genotype or haplotype frequencies and risk of AD. Further, there was no effect of genotype on platelet membrane beta-secretase activity. We conclude that common or potentially functional genetic variation in these BACE1 interacting proteins does not affect platelet membrane beta-secretase activity or contribute to risk of AD in this population.

Rapid Surface Plasmon Resonance Immunoassay for the Determination of Deoxynivalenol in Wheat, Wheat Products, and Maize-Based Baby Food

A rapid screening assay (9 min/sample) has been developed and validated for the detection of deoxynivalenol in durum wheat, wheat products, and maize-based baby foods using an SPA biosensor. Through a single laboratory validation, the limits of detection (LOD) for wheat, wheat-based breakfast cereal, and maize-based baby food were 57, 9, and 6 mu g/kg, respectively. Intra-assay and interassay precisions were calculated for each matrix at the maximum and half-maximum European Union regulatory limits and expressed as the coefficient of variation (CV). All CVs fell below 10% with the exception of the between-run CV for breakfast cereal. Recoveries at the concentrations tested ranged from 92 to 115% for all matrices. Action limits of 161, 348, and 1378 mu g/kg were calculated for baby food, wheat-based breakfast cereal, and wheat, respectively, and the linear range of the assay was determined as 250-2000 mu g/kg.

Hapten synthesis and antibody production for the development of a melamine immunoassay

The incorporation of melamine into food products is banned but its misuse has been widely reported in both animal feeds and food. The development of a rapid screening immunoassay for monitoring of the substance is an urgent requirement. Two haptens of melamine were synthesized by introducing spacer arms of different lengths and structures on the triazine ring of the analyte molecular structure. 6-Aminocaproic acid and 3-mercaptopropionic acid were reacted with 2-chloro-4,6-diamino-1,3,5-triazine (CAAT) to produce hapten 1[3-(4,6-diamino-1,6-dihydro-1,3,5-triazin-2-ylamino) hexanoic acid] and hapten 2[3-(4,6-diamino-1,6-dihydro-1,3,5-triazin-2-ylthio) propanoic acid]. respectively. The molecular structures of the two haptens were identified by I H nuclear magnetic resonance spectrometry, mass spectrometry and infrared spectrometry. An immunogen was prepared by coupling hapten 1 to bovine serum albumin (BSA). Two plate coating antigens were prepared by coupling both haptens to egg ovalbumin (OVA). A competitive indirect enzyme-linked immunosorbent assay (ciELISA) was developed to evaluate homogeneous and heterogeneous assay formats. The results showed that polyclonal antibodies with high titers were obtained, and the heterogeneous immunoassay format demonstrated a better performance with an IC50 of 70.6 ng mL(-1), a LOD of 2.6 ng mL(-1) and a LOQ of 7.6 ng mL(-1). Except for cyromazine, no obvious cross-reactivity to common compounds was found. The data showed that the hapten synthesis was successful and the resultant antisera could be used in an immunoassay for the rapid and sensitive detection of this banned chemical. (C) 2010 Elsevier B.V. All rights reserved.

A rapid optical immunoassay for the screening of T-2 and HT-2 toxin in cereals and maize-based baby food

A rapid surface plasmon resonance (SPR) screening assay has been developed for the combined detection of T-2 and HT-2 toxins in naturally contaminated cereals using a sensor chip coated with an HT-2 toxin derivative and a monoclonal antibody. The antibody raised against HT-2 displayed high cross-reactivity with T-2 toxin while there was no cross-reaction observed with other commonly occurring trichothecenes. A simple extraction procedure using 40% methanol was applied to baby food, breakfast cereal, and wheat samples prior to biosensor analysis. Limits of detection (LOD) for each matrix were determined as 25 mu g kg(-1) for baby food and breakfast cereal and 26 mu g kg(-1) for wheat. Intra-assay precision (n = 6) was calculated for each matrix. The results were expressed as the relative standard deviation and determined as 2.8% (100 mu g kg(-1)) and 1.8% (200 mu g kg(-1)) in breakfast cereal, 4.6% (50 mu g kg(-1)) and 3.6% (100 mu g kg(-1)) in wheat and 0.97% (25 mu g kg(-1)) and 6.3% (50 mu g kg(-1)) in baby food. Between run precision (n = 3) performed at the same levels yielded relative standard deviations of 6.7% and 3.9% for breakfast cereals, 3.3% and 1.6% for wheat and 6.8% and 0.08% for baby food, respectively. (C) 2010 Elsevier B.V. All rights reserved.

Significant correlation in linkage signals from genome-wide scans of schizophrenia and schizotypy

Prior family and adoption studies have suggested a genetic relationship between schizophrenia and schizotypy. However, this has never been verified using linkage methods. We therefore attempted to test for a correlation in linkage signals from genome-wide scans of schizophrenia and schizotypy. The Irish study of high-density schizophrenia families comprises 270 families with at least two members with schizophrenia or poor-outcome schizoaffective disorder (n = 637). Non-psychotic relatives were assessed using the structured interview for schizotypy (n = 746). A 10-cM multipoint, non-parametric, autosomal genomewide scan of schizophrenia was performed in Merlin. A scan of a quantitative trait comprising ratings of DSM-III-R criteria for schizotypal personality disorder in non-psychotic relatives was also performed. Schizotypy logarithm of the odds (LOD) scores were regressed onto schizophrenia LOD scores at all loci, with adjustment for spatial autocorrelation. To assess empirical significance, this was also carried out using 1000 null scans of schizotypy. The number of jointly linked loci in the real data was compared to distribution of jointly linked loci in the null scans. No markers were suggestively linked to schizotypy based on strict Lander Kruglyak criteria. Schizotypy LODs predicted schizophrenia LODs above chance expectation genome wide (empirical P = 0.04). Two and four loci yielded nonparametric LOD (NPLs) > 1.0 and > 0.75, respectively, for both schizophrenia and schizotypy (genome-wide empirical P = 0.04 and 0.02, respectively). These results suggest that at least a subset of schizophrenia susceptibility genes also affects schizotypy in non-psychotic relatives. Power may therefore be increased in molecular genetic studies of schizophrenia if they incorporate measures of schizotypy in non-psychotic relatives.