罗氏沼虾淀粉酶基因克隆及其表达规律分析

【目的】克隆罗氏沼虾（Macrobrachium rosenbergii）淀粉酶基因（AMY）,并进行生物学信息分析及其表达规律研究,为揭示AMY基因的生物学功能及其对罗氏沼虾生长发育的调控作用机理提供理论依据。【方法】PCR克隆罗氏沼虾AMY基因编码区（CDS）序列,利用ExPASy ProtParam、ExPASy ProtScale、SignalP-5.0、MegAlign及Lasergenev8.0等在线软件进行生物信息学分析,应用实时荧光定量PCR检测AMY基因在罗氏沼虾不同组织（腹神经节、胃、心脏、鳃组织、性腺、肝胰腺、肌肉和肠道）中的表达情况,并明确其在生长快速家系（FG）和生长慢速家系（SG）个体肌肉中的表达模式。【结果】罗氏沼虾AMY基因CDS序列全长2121 bp,共编码706个氨基酸残基;其编码蛋白分子量为76.87 kD,理论等电点（pI）为4.63,属于不稳定的亲水性蛋白,包含2个典型的淀粉酶结构域;在罗氏沼虾AMY蛋白二级结构中,α-螺旋占13.88%,延伸链占21.39%,无规则卷曲占64.73%。罗氏沼虾AMY氨基酸序列与克氏原螯虾AMY氨基酸序列的相似性最高（62.6%）,与大珠母贝AMY氨基酸序列的相似性较低（48.9%）;基于AMY氨基酸序列相似性构建的系统发育进化树显示,罗氏沼虾与克氏原螯虾的亲缘关系最近,而与中华绒螯蟹的亲缘关系较远。AMY基因在罗氏沼虾不同组织中广泛表达,但存在性别差异性和组织特异性,具体表现为:雌性个体肠道中的相对表达量极显著高于雄性个体（P<0.01,下同）,鳃组织中的相对表达量显著高于雄性个体（P<0.05）,而肝胰腺中的相对表达量极显著低于雄性个体。罗氏沼虾AMY基因在FG个体肌肉中的相对表达量极显著高于SG个体。【结论】AMY基因在罗氏沼虾的生长发育过程中发挥重要作用,广泛表达于罗氏沼虾的不同组织中,但存在性别差异性和组织特异性。     

Germ line p53 mutations in a familial syndrome of breast cancer, sarcomas, and other neoplasms

Familial cancer syndromes have helped to define the role of tumor suppressor genes in the development of cancer. The dominantly inherited Li-Fraumeni syndrome (LFS) is of particular interest because of the diversity of childhood and adult tumors that occur in affected individuals. The rarity and high mortality of LFS precluded formal linkage analysis. The alternative approach was to select the most plausible candidate gene. The tumor suppressor gene, p53, was studied because of previous indications that this gene is inactivated in the sporadic (nonfamilial) forms of most cancers that are associated with LFS. Germ line p53 mutations have been detected in all five LFS families analyzed. These mutations do not produce amounts of mutant p53 protein expected to exert a trans-dominant loss of function effect on wild-type p53 protein. The frequency of germ line p53 mutations can now be examined in additional families with LFS, and in other cancer patients and families with clinical features that might be attributed to the mutation.     

Human hair growth deficiency is linked to a genetic defect in the phospholipase gene LIPH

The molecular mechanisms controlling human hair growth and scalp hair loss are poorly understood. By screening about 350,000 individuals in two populations from the Volga-Ural region of Russia, we identified a gene mutation in families who show an inherited form of hair loss and a hair growth defect. Affected individuals were homozygous for a deletion in the LIPH gene on chromosome 3q27, caused by short interspersed nuclear element-retrotransposon-mediated recombination. The LIPH gene is expressed in hair follicles and encodes a phospholipase called lipase H (alternatively known as membrane-associated phosphatidic acid-selective phospholipase A1alpha), an enzyme that regulates the production of bioactive lipids. These results suggest that lipase H participates in hair growth and development.     

葡萄风信子黄酮醇合酶基因克隆和表达分析

以葡萄风信子‘亚美尼亚’的根、茎、叶、花为材料,采用RT-PCR技术克隆得到一条FLS基因,命名为MaFLS2。生物信息学分析表明,该基因cDNA全长为1 152 bp,含有完整的开放读码框1 056 bp,编码353个氨基酸,MaFLS2蛋白是一种亲水性蛋白,氨基酸序列含有典型的DIOX_N结构域(2-酮戊二酸-双加氧酶活性的N端区域)和2OG-FeII_Oxy超家族结构域,属于2-酮戊二酸-双加氧酶(2-ODDs)蛋白家族。实时定量PCR结果显示,MaFLS2基因在蓝白葡萄风信子中的表达存在组织特异性,蓝色品种‘亚美尼亚’中在花蕾未着色时表达量最高,而白色品种‘白丽人’在根中表达量最高, 在花发育的不同阶段,MaFLS2基因在‘亚美尼亚’中的表达峰值出现在花蕾未着色时期,基因表达总体呈现出先降低后升高再降低的趋势,而在‘白丽人’中的表达峰值出现在转色中期,总体呈现出先升后降的趋势。研究结果为开展单子叶植物花色育种提供了理论依据。     

原发性开角型青光眼中医证型、证候、证素文献分析研究

目的 通过文献研究,分析原发性开角型青光眼（primary open angle glaucoma,POAG）证型分布特点及其证候、证素特点,为临床POAG的诊治提供依据。方法 检索CNKI（中国知网数据库）、VIP（维普数据库）、CBM（中国生物医学文献数据库）和Wanfang数据库（万方数据库）,采用EpiData3.1软件建立POAG证候文献数据库,应用统计软件SPSS 19.0进行统计分析。结果 最终纳入文献11篇,包括644例不同证型的患者,规范后的POAG证型11种,最常见的4种证型为：肝肾亏虚证（26.55%）、肝郁化火证（23.45%）、痰湿泛目证（18.94%）、肝郁气滞证（13.20%）,与POAG相关的病位证素为肝,病性证素主要与阴虚、血虚、阳亢、气滞相关。结论 本研究初步反映了POAG常见证型、证候分布特点,并对其证素特点进行了概括,为临床POAG的诊疗提供依据。     

猪传染性胃肠炎病毒纤突蛋白基因的修饰及原核表达的研究

猪传染性胃肠炎病毒纤突蛋白是诱导猪产生特异性中和抗体的主要结构蛋白,包括A,B,C,D4个抗原决定位。为进一步生产TGEV植物转基因疫苗提供可操作性强、具免疫活性的外源基因,以猪传染性胃肠炎华毒弱株(TGEVH)的S基因为基础,扩增并连接4个抗原决定位相应的碱基序列,构建pPROEX-HTa原核表达载体,转化大肠杆菌诱导表达。结果表明,修饰后的S蛋白的表达量为19.8%,仍能和相应的血清发生抗原抗体反应。     

Pht1家族在几种植物中的研究进展

植物磷转运蛋白是植物磷营养中必需的一种膜蛋白。植物磷转运子在植物根系中负责磷的吸收、转运,其表达受磷调控,磷元素广泛存在于动植物组织中,是植物生长所必需的大量无机营养元素之一,在诸多代谢过程中都起着举足轻重的作用。在植株中,磷素通过磷酸盐转运蛋白吸收和转运,该蛋白在调控植株对磷素吸收、利用效率等方面具有重要作用。植物基因组中含有大量推测的以基因家族的形式存在的编码磷转运蛋白基因。目前已知的磷转运子分为五大家族Pht1、Pht2、Pht3、Pho1和Pho2。文中主要综述了水稻、大豆、玉米、小麦、拟南芥、番茄、苜蓿、马铃薯中Pht1家族的结构、功能及表达调控方面的研究进展。     

Genome of the host-cell transforming parasite Theileria annulata compared with T. parva

Theileria annulata and T. parva are closely related protozoan parasites that cause lymphoproliferative diseases of cattle. We sequenced the genome of T. annulata and compared it with that of T. parva to understand the mechanisms underlying transformation and tropism. Despite high conservation of gene sequences and synteny, the analysis reveals unequally expanded gene families and species-specific genes. We also identify divergent families of putative secreted polypeptides that may reduce immune recognition, candidate regulators of host-cell transformation, and a Theileria-specific protein domain [frequently associated in Theileria (FAINT)] present in a large number of secreted proteins.     

黑松冠层节肢动物群落的组成与结构

2006年5-9月,采用枝条套袋取样技术对浙江省舟山市松材线虫病Bursaphelenchus xylophilus疫区内黑松Pinus thunbergii冠层节肢动物群落的组成和结构进行了研究。结果表明：供调查的黑松冠层中,共有节肢动物12目45科95种,共计3 077个个体。其中捕食性昆虫类群有10科17种89头,食叶害虫类群11科16种1 477个,枝梢害虫类群8科18种1 186个,蜘蛛类群11科33种287只,游走性类群5科13种38个。主要害虫种类有浙江黑松叶蜂Nesodiprion zhejiangensis,草履蚧Drosicha corpulenta,松沫蝉 Aphrophora flavipes,松褐天牛 Monochamus alternatus等。各功能集团主要分布在树冠的中下层,除捕食性昆虫外,总群落及其他各类群对方位的选择性较小,捕食性昆虫主要分布在向阳的东部、南部和西部。以食叶害虫和枝梢害虫为主的害虫类群在5月份的优势集中性最高,在7月物种丰富度最高,6－8月危害最为严重。而蜘蛛类和捕食性昆虫等捕食性类群在调查的各个月份中的优势集中性均比较稳定,仅在8月有所降低。图3表1参23     

Common sequence polymorphisms shaping genetic diversity in Arabidopsis thaliana

The genomes of individuals from the same species vary in sequence as a result of different evolutionary processes. To examine the patterns of, and the forces shaping, sequence variation in Arabidopsis thaliana, we performed high-density array resequencing of 20 diverse strains (accessions). More than 1 million nonredundant single-nucleotide polymorphisms (SNPs) were identified at moderate false discovery rates (FDRs), and approximately 4% of the genome was identified as being highly dissimilar or deleted relative to the reference genome sequence. Patterns of polymorphism are highly nonrandom among gene families, with genes mediating interaction with the biotic environment having exceptional polymorphism levels. At the chromosomal scale, regional variation in polymorphism was readily apparent. A scan for recent selective sweeps revealed several candidate regions, including a notable example in which almost all variation was removed in a 500-kilobase window. Analyzing the polymorphisms we describe in larger sets of accessions will enable a detailed understanding of forces shaping population-wide sequence variation in A. thaliana.     

鹌鹑piRNAs和Piwil1基因克隆及表达研究

 【目的】目前,国内外尚无有关禽类piRNAs及其结合蛋白Piwi的研究报道,开展禽类小分子RNA及其结合蛋白的研究将为小分子RNA的遗传特性、发生和消失机理以及转基因鸡等研究提供基础依据。【方法】【结果】本研究以鹌鹑为研究对象,通过构建cDNA文库和TA克隆测序的方法首次从睾丸组织中获得了13个piRNAs克隆序列和Piwil1基因的CDS序列,同时采用Q-PCR技术分析了鹌鹑不同生命阶段不同组织中piRNAs和Piwil1基因的表达规律。【结论】结果表明,鹌鹑piRNAs长度与哺乳动物相似;Piwil1基因与红色原鸡有较高的同源性,包含PAZ和Piwi结构域,无信号肽剪切位点,推测其属于Piwi家族蛋白,可能通过降低结合RNA的能力或者影响其他转录因子与结构基因启动子的亲和性发挥调控作用;在睾丸组织中,piRNAs和Piwil1蛋白的表达量均是随着个体的不断发育而增加,推断在禽类中piRNAs可能与Piwi蛋白相互作用共同调控精子的发生过程。     

淡水养殖凡纳滨对虾生长相关基因HDAC1克隆及其表达分析

【目的】分析组蛋白去乙酰基酶1基因（HDAC1）在不同淡水生长特性家系凡纳滨对虾中的表达情况,明确HDAC1基因在对虾淡水养殖过程中的功能作用,为揭示凡纳滨对虾生长调控的分子机制提供参考依据。【方法】通过RACE克隆凡纳滨对虾HADC1基因cDNA序列,利用DNAMAN、ProtParam、ProtScale、PSOR...     

The protein kinase complement of the human genome

We have catalogued the protein kinase complement of the human genome (the "kinome") using public and proprietary genomic, complementary DNA, and expressed sequence tag (EST) sequences. This provides a starting point for comprehensive analysis of protein phosphorylation in normal and disease states, as well as a detailed view of the current state of human genome analysis through a focus on one large gene family. We identify 518 putative protein kinase genes, of which 71 have not previously been reported or described as kinases, and we extend or correct the protein sequences of 56 more kinases. New genes include members of well-studied families as well as previously unidentified families, some of which are conserved in model organisms. Classification and comparison with model organism kinomes identified orthologous groups and highlighted expansions specific to human and other lineages. We also identified 106 protein kinase pseudogenes. Chromosomal mapping revealed several small clusters of kinase genes and revealed that 244 kinases map to disease loci or cancer amplicons.     

基于AFLP标记的不同群体厚朴遗传多样性分析

[目的]探讨厚朴种源间、种源内家系及家系内个体间的遗传多样性,为厚朴良种选育提供理论依据。[方法]采用AFLP分子标记技术,分析来自湖北、广西和浙江的厚朴种源间、半同胞家系间及家系内单株间的遗传多样性。[结果]3个种源材料遗传距离为0.048 2~0.128 4,广西资源和浙江景宁2个群体相似性较高。种群遗传多样性平均水平、总种群基因多样度、基因流分别为0.203 5、0.254 1、3.421 7。观测等位基因数和有效等位基因数平均数分别为1.658 3和1.341 4;Nei’s基因多样性和Shannon多样性指数均值分别为0.203 5和0.308 7,其中湖北五峰的遗传多样性水平最高(为0.225 8),浙江景宁遗传多样性水平最低(为0.173 9)。湖北五峰、广西资源、浙江景宁家系内遗传多样性分别为0.264 4、0.181 9、0.173 8,Shannon多样性指数为0.415 4。[结论]湖北五峰种源遗传多样性水平最高,湖北五峰家系内的遗传多样性最丰富,存在较大的遗传分化。不同种源内家系之间的多样性指数及信息指数相差不多,说明在各自的生态环境中的变异水平相近。在厚朴良种选育过程中应充分重视优良种源、优良家系与优良单株的配合选择。     

建鲤IGFⅠa基因的SNPs位点筛选及其与增重的相关性分析

机体生长主要受到促生长激素释放激素—生长激素—胰岛素样生长因子(GHRH-GH-IGFs)生长轴调控,类胰岛素生长因子IGFⅠ是生长激素（GH）发挥生物学功能的重要传导因子。实验特异性扩增了建鲤(Cyprinus carpio var. jian) IGFⅠa基因的5个外显子和3个内含子（内含子1、内含子3和内含子4）。通过比对10尾建鲤的序列,共找到SNPs位点8个。使用PCRRFLP方法检测了5个家系共372尾建鲤的内含子1_C175G,12个家系共987尾的内含子1_A993G和内含子4_A511C 3个位点。内含子1_C175G雌雄个体均以GG型频率为最高,分别是0.44和0.43;此位点在幼鱼和成鱼时期与雌、雄建鲤增重均无相关性;内含子1_A993G雌雄个体均以AG型频率最高,分别是0.76和0.72;此位点在成鱼阶段与雄性建鲤增重呈显著相关(P<0.05);内含子4_A511C在雌雄个体中均以CC型频率最高,分别是0.48和0.47;其在幼鱼和成鱼阶段均与增重存在极显著的相关性(P<0.01)。本次试验表明在建鲤体内,IGFⅠ基因在不同生长阶段的表达量不同,且处于同一生长阶段的雌雄个体间的表达也存在差异。内含子1_A993G、内含子4_A511C均与建鲤增重存在相关性,可以考虑作为建鲤分子育种的相关依据。     

A previously undetected MHC gene with an unusual periodic structure

The major histocompatibility complex is a chromosomal segment embodying several gene clusters among which those with immune functions are the best characterized. This region is suspected to host other as yet undetected genes whose characterization may shed light on the population genetics and evolution of the whole gene complex and thus on its unexplained character of marker locus for a number of diseases of nonimmune or unknown pathogenesis. A novel gene was identified that is transcribed in all tissues tested and is located in mouse and man between the CA and Bf genes of the H-2 and HLA complexes, respectively. From the nucleotide sequence, derived from liver complementary DNA clones, it is predicted that this novel single-copy gene encodes a 42-kilodalton polypeptide that bears no recognizable relation to the protein families known so far, but it displays striking hallmarks of natural selection.     

Detection of TGEV Antibody by Enzyme-Linked Immunosorbent Assay Using Recombinant Nucleocapsid Proteins

An enzyme linked immunosorbent assays (ELISA) based on recombinant nucleocapsid (N) protein generated in Escherichia coli was evaluated for its sensitivity and specificity for diagnosis of transmissible gastroenteritis virus (TGEV) infection.The N gene encoding the N protein was cloned and expressed as a fusion protein with His tag protein in E. coli. The recombinant N protein migrated at 42 kDa and reacted with His6 tag specific monoclonal antibody by immunoblotting.Recombinant N protein ELISA (rnELISA) demonstrated 97.5% specificity among 80 TGEV-free individuals, and 97.3%sensitivity ranging among 110 clinical samples with TGEV. Taken together, these results indicated that nucleocapsid may be a useful antigen for the sera-diagnosis of TGEV and it was also suggested that the ELISA is a highly sensitive and specific test for detecting antibodies against TGEV.     

河北小五台山半翅目昆虫的多样性研究

[目的]探明河北小五台自然保护区的蝽类物种及其组成情况。[方法]2009年在河北小五台山选取5条不同样带,对半翅目昆虫群落组成和多样性进行研究。[结果]共采集标本3246只,隶属于21科98属145种,其中蝽科最多,占43．31％;蝽科、缘蝽科、同蝽科、姬缘蝽科和盲蝽科共同构成该地区的优势类群,这5个科的个体数量占总采集数量的76．59％。半翅目群落种一多度关系接近于正态分布模型。涿鹿杨家坪的物种多样性指数、物种数和个体数量最高;蔚县王喜洞的物种多样性指数、物种数和个体数量最低。聚类分析结果表明蔚县小五台、蔚县山涧口和蔚县王喜洞半翅目昆虫群落相似度较高,蔚县金河口和涿鹿杨家坪的相似度较高。[结论]该研究为自然保护区昆虫种质资源的保护提供参考。