Ontogeny of odorant receptor gene expression in zebrafish,Danio rerio

We cloned three putative odorant receptor (OR) genes from the zebrafish to use as in situ hybridization probes to follow the temporal patterns of neurons expressing OR genes through a developmental progression from embryo (12 h postfertilization) to adult. The identification of these genes is supported by sequence homology to previously reported ORs and by the morphology and location of labeled cells in in situ hybridization experiments. Cells expressing OR mRNA were first observed in the olfactory placodes between 31 and 38 h after fertilization (fish reared at 26°C). Initially, only single cells were observed to hybridize the probe; the number of labeled cells increased throughout the remainder of embryogenesis and through postembryonic growth and morphogenesis of the olfactory organ. At all ages, the positively hybridizing cells were scattered throughout the olfactory epithelium but not in the nonsensory epithelium of the olfactory organ. © 1996 John Wiley & Sons, Inc.     

Natural hybridization of two species of tetraploid barbels: Barbus meridionalis and Barbus barbus (Osteichtyes, Cyprinidae) in southern France

Natural hybridization of Barbus barbus and B. meridionalis has been demonstrated in southern France. A genetical study (isozyme electrophoresis) of these species and their hybrids revealed the characteristics of hybridization. Ten enzyme markers and five morphological parameters were used to distinguish between the two parent species. Enzymatic divergence between the parent species included fixed allelic differences at certain loci and reduction in enzymatic activity, including the silencing of certain genes. A morphological study revealed a good correlation between the isozyme markers and morphological characters. Backcrosses were observed and this raises the question of the integrity of the parent species.     

飞蝗发育相关基因Omb的克隆、原核表达及时空表达分析

【目的】本研究克隆飞蝗Locusta migratoria发育相关的optomotor-blind(Omb)基因,并对其进行序列和表达分析,旨在更好地了解Omb基因在飞蝗翅发育中的作用及为进一步蝗灾的治理和防治提供新的理论依据。【方法】利用RT-RCR扩增飞蝗Omb基因cDNA序列,采用生物信息学软件分析该基因的核苷酸和氨基酸序列,利用MEGA 6. 0构建昆虫纲分子系统进化树;构建重组表达载体pET-30a/LmOmb,转化到大肠杆菌Escherichia coli BL21(DE3)中,SDS-PAGE及Western blot鉴定重组表达蛋白;基于qPCR技术分析Omb基因在飞蝗不同发育时期及成虫不同组织中的表达谱。【结果】克隆获得飞蝗Omb基因部分cDNA序列,命名为LmOmb(GenBank登录号:MG867658),其长792 bp,编码264个氨基酸,在第37-219位氨基酸之间存在一个T-box superfamily保守结构域。同源序列比对分析表明Lm Omb与褐飞虱Nilaparvata lugens Nl Omb氨基酸序列一致性为93%。在IPTG诱导下目标蛋白以6×His标签融合蛋白的形式在宿主菌中得到稳定表达。荧光定量PCR结果显示LmOmb基因在飞蝗不同发育时期均有表达,其中胚胎期的表达量最高,进入若虫期后表达量下降,且各龄若虫之间表达量相对平稳。Lm Omb基因在雌性和雄性成虫的胸部、足、腹部和翅中都有表达,且雌性和雄性之间表达量明显不同。【结论】LmOmb基因可能参与了飞蝗的胚胎发育。研究结果为进一步研究飞蝗LmOmb的功能提供了依据。     

Regulation of gonadotropin-releasing hormone (GnRH) gene expression in hypothalamic neuronal cells

Summary 1. Gonadotropin-releasing hormone (GnRH) is the hypothalamic releasing factor that controls pituitary gonadotropin subunit gene expression and indirectly gametogenesis and steroidogenesis from the gonad, which results in reproductive competence.2. GnRH is synthesized in only about 1000 neurons in the hypothalamus and released in an episodic fashion down the median eminence to regulate gonadotropin biosynthesis.3. Although much is known about the secretory dynamics of GnRH release, little is known about the pretranslational control of GnRH biosynthesis due to lack of appropriate model systems. The recent availability of immortalized neuronal cell lines that produce GnRH allows investigators for the first time to begin to dissect the factors that directly regulate GnRH gene expression.4. This article reviews the current state of knowledge concerning the mechanisms that direct tissue-specific and peptide hormone control of GnRH biosynthesis.     

Oligoryzomys flavescens (Rodentia, Muridae): gene flow among populations from central-eastern Argentina

In species acting as hosts of infectious agents, the extent of gene flow between populations is of particular interest because the expansion of different infectious diseases is usually related to the dispersal of the host. We have estimated levels of gene flow among populations of the sigmodontine rodent Oligoryzomys flavescens, in which high titers of antibodies have been detected for a Hantavirus in Argentina that produces a severe pulmonary syndrome. Enzyme polymorphism was studied by means of starch gel electrophoresis in 10 populations from the area where human cases of Hantavirus have occurred. Genetic differentiation between populations was calculated from F_ST values with the equation Nm = [(1/F_ST−1]/4. To assess the relative importance of current gene flow and historical associations between populations, the relationship of population pairwise log Nm and log geographic distance was examined. Low F_ST (mean = 0.038) and high Nm (15.27) values suggest high levels of gene flow among populations. The lack of an isolation by distance pattern would indicate that this species has recently colonized the area. The northernmost population, located on the margin of a great river, shows very high levels of gene flow with the downstream populations despite the large geographic distances. Passive transport of animals down the river by floating plants would promote unidirectional gene flow. This fact and the highest mean heterozygosity of that northernmost population suggest it is a center of dispersal within the species' range. This revised version was published online in July 2006 with corrections to the Cover Date.     

棉卷叶野螟泛素基因的克隆、序列分析及原核表达

本研究用RT-PCR方法,克隆了棉卷叶野螟Haritalodes derogata (Fabricius)泛素基因编码区,GenBank登录号为EU580145。序列分析表明,该编码区长228 bp,编码76个氨基酸,推测的编码蛋白的相对分子质量和等电点分别为8.53 kD和5.83。同源性比较发现,棉卷叶野螟泛素基因与其他10种昆虫泛素基因在氨基酸水平上具有93%以上的相似性。系统发育树显示棉卷叶野螟与斜纹夜蛾Spodoptera litura (Fabricius)遗传距离较近,通过同源建模获得了该棉卷叶野螟基因编码蛋白的理论三维结构。将棉卷叶野螟泛素基因与pET-32a(+)连接,构建原核表达载体pET-32a-ub,经IPTG诱导,棉卷叶野螟泛素基因在大肠杆菌BL21(DE3) 中高效表达。本研究成功克隆了棉卷叶野螟泛素基因的编码区,并经Western blotting分析证明实现了该基因的原核表达,为进一步研究其在该昆虫体内的作用机理奠定了基础。     

Tissue-specific gene expression in soybean (Glycine max) detected by cDNA microarray analysis

We have constructed cDNA microarrays for soybean (Glycine maxL. Merrill), containing approximately 4,100 Unigene ESTs derived from axenic roots, to evaluate their application and utility for functional genomics of organ differentiation in legumes. We assessed microarray technology by conducting studies to evaluate the accuracy of microarray data and have found them to be both reliable and reproducible in repeat hybridisations. Several ESTs showed high levels (50 fold) of differential expression in either root or shoot tissue of soybean. A small number of physiologically interesting, and differentially expressed sequences found by microarray analysis were verified by both quantitative real-time RT-PCR and Northern blot analysis. There was a linear correlation (r² = 0.99, over 5 orders of magnitude) between microarray and quantitative real-time RT-PCR data. Microarray analysis of soybean has enormous potential not only for the discovery of new genes involved in tissue differentiation and function, but also to study the expression of previously characterised genes, gene networks and gene interactions in wild-type, mutant or transgenic plants.     

婚飞行为影响中华蜜蜂性成熟处女蜂王的基因表达

婚飞是性成熟处女蜂王与雄蜂交配过程中的一个重要前奏, 在该过程中蜂王体内伴随着一系列重要的生理变化。为了探究中华蜜蜂Apis cerana cerana处女蜂王婚飞过程中基因表达变化, 本研究利用数字基因表达谱(digital gene expression, DGE) 技术分析了中华蜜蜂性成熟处女蜂王飞行与未飞行之间的基因表达差异。经DGE测序, 分别从两个样品中获得5.98和6.01 百万条Clean标签。通过分析检测到250个基因有差异表达, 其中133个基因在飞行蜂王中上调表达, 117个基因在飞行蜂王中下调表达。这些差异基因可以归类到348个功能性类别和142个生化途径。结果表明中华蜜蜂性成熟处女蜂王在婚飞过程中大量基因的表达发生了变化。这些结果为进一步研究中华蜜蜂蜂王婚飞过程中生理变化的分子机制提供了重要的基因表达信息。     

High level of gene silencing in the tetraploid goldfish

The goldfish, Carassius auratus, is a karyotypically tetraploid form that expresses only 19% of its enzyme-encoding loci in duplicate. This level of gene duplication is among the lowest reported among tetraploid cypriniform fishes and may be related to the intense selective and drift processes associated with its domestication.     

扶桑绵粉蚧组织蛋白酶B基因的克隆、原核表达和不同发育阶段表达分析

昆虫组织蛋白酶B在昆虫代谢过程中发挥重要作用。本研究利用RACE技术克隆了扶桑绵粉蚧Phenacoccus solenopsis Tinsley组织蛋白酶B基因的开放阅读框（ORF）序列, 命名为PsCb （GenBank登录号: JQ727999）。生物信息学分析表明, 该基因的开放阅读框包含927 bp的片段, 编码308个氨基酸。多序列比对表明, 该基因编码的蛋白在N端变异较大, 在C端保守性高。组织蛋白酶B基因的系统进化树结果表明扶桑绵粉蚧组织蛋白酶独自成为一支。原核表达电泳检测到一条大约35 kDa的目的条带, 与预测的蛋白分子量相符。组织蛋白酶B基因在扶桑绵粉蚧各个虫态均有表达, 卵期表达量相对较低, 2龄若虫期达到最高峰, 然后下降。本研究为进一步研究该基因的功能并开发出组织蛋白酶抑制剂, 从而研制出扶桑绵粉蚧杀卵剂和胚胎发育抑制剂等提供理论依据。     

Evaluation of deleted in malignant brain tumors 1 (DMBT1) gene expression in bladder carcinoma cases: preliminary study

This study was undertaken to evaluate the expression of DMBT1 in bladder cancer and its correlation with clinico-pathological parameters analyzed in bladder carcinoma patients. We investigated DMBT1 in 56 paraffin embedded specimens of transitional cell carcinoma of the urinary bladder. We assessed DMBT1 gene expression at mRNA level by RT-PCR. Our results show 100% expression of DMBT1 in bladder carcinoma samples. Due to this preliminary results; gene expression was compared to tumor grade, and a significant difference was detected between grade 1 and 3 (p?=?0.028). The down-regulation of DMBT1 gene expression in carcinomas suggests the possible role in bladder cancer.     

真核基因的快速克隆及表达

以细胞间隙连接蛋白基因Cx26作为目的基因,通过T-A载体介导,构建真核表达重组载体pcDNA3.1( ) /Cx26,重组表达载体转染人鼻咽癌细胞株HNE1,表达Cx26间隙连接蛋白。     

中华蜜蜂OBP3基因的克隆、原核表达及组织表达谱

【目的】气味结合蛋白质（odorant binding proteins, OBPs）参与气味分子的识别,在蜜蜂嗅觉中扮演重要的角色。本研究旨在克隆中华蜜蜂 Apis cerana cerana OBP3基因,以制备多克隆抗体。【方法】运用RT-PCR技术从中华蜜蜂头部总RNA中扩增OBP3基因,将该基因亚克隆入原核表达载体pET-28a并转入大肠杆菌Escherichia coli Rosetta (DE3)中诱导表达获得融合蛋白质,融合蛋白质经纯化后免疫新西兰白兔制备多克隆抗体,最后分别用间接ELISA和Western Blot检测抗体的效价和特异性,并采用荧光定量PCR检测OBP3基因在中蜂不同组织中的表达。【结果】克隆得到了中华蜜蜂OBP3基因AccOBP3（GenBank登录号KJ026357）,大小为444 bp。 SDS-PAGE结果显示融合蛋白成功表达。制备的多克隆抗体效价高于1∶40 000,且具有很高的特异性。荧光定量PCR结果表明,AccOBP3基因在腿部和触角中显著高表达（P<0.01）,胸部中次之（P<0.01）,头部和腹部中显著低表达,后两者表达量差异不显著（P>0.05）。【结论】OBP3基因在中蜂触角有高转录活性。本研究实现了中蜂OBP3基因的原核表达,并制备了兔抗中蜂OBP3多克隆抗体,为深入研究中蜂OBP3基因的功能奠定基础 。     

西方蜜蜂发育基因AmWnt1的克隆鉴定及时空表达分析

本研究旨在克隆鉴定西方蜜蜂Apis mellifera发育相关基因AmWnt1,分析其在不同发育时期和刚出房工蜂不同组织的表达特征,为进一步研究Wnt1基因功能提供理论参考。根据NCBI中AmWnt1基因序列信息,利用Primer 6.0设计引物,RT-PCR扩增AmWnt1基因完整的CDS序列,进行生物信息学预测,用推导的氨基酸序列构建系统进化树;利用荧光定量PCR检测该基因在卵（1日龄、2日龄和3日龄）、幼虫（1日龄、3日龄和5日龄）、预蛹（1日龄和3日龄）、蛹（0日龄、2日龄、4日龄、6日龄和8日龄）、刚出房工蜂、哺育蜂和采集蜂以及刚出房工蜂8个组织中相对表达量。克隆获得西方蜜蜂的Wnt1基因CDS序列,命名为AmWnt1,上传NCBI,获得GenBank登录号MT993937。全长1 239 bp,编码412个氨基酸,预测等电点为9.48,相对分子质量为46.40313 kDa。序列比对和系统进化树结果表明：AmWnt1蛋白与其它膜翅目昆虫聚为一类,其中和东方蜜蜂Apis cerana亲缘关系最近,序列相似度为99.50%。时空表达谱结果表明：AmWnt1基因在各个时期中均有表达,且在胚胎后期表达量最高,预蛹期和蛹前期表达量相对较高,其它时期表达量相对较低;AmWnt1基因在刚出房工蜂头、胸、触角表达量高于其它组织。AmWnt1可能参与西方蜜蜂胚胎晚期的神经系统发育和化蛹过程中的四肢发育等关键历程,为进一步研究AmWnt1功能提供了理论参考。     

Identification of reproductive sex-biased gene expression in Asparagopsis taxiformis (lineage 6) gametophytes

The sub-tropical red seaweed Asparagopsis taxiformis is of significant interest due to its ability to store halogenated compounds, including bromoform, which can mitigate methane production in ruminants. Significant scale-up of aquaculture production of this seaweed is required; however, relatively little is known about the molecular mechanisms that control fundamental physiological processes, including the regulatory factors that determine sexual dimorphism in gametophytes. In this study, we used comparative RNA-sequencing analysis between different morphological parts of mature male and female A. taxiformis (lineage 6) gametophytes that resulted in greater number of sex-biased gene expression in tips (containing the reproductive structures for both sexes), compared with the somatic main axis and rhizomes. Further comparative RNA-seq against immature tips was used to identify 62 reproductive sex-biased genes (59 male-biased, 3 female-biased). Of the reproductive male-biased genes, 46% had an unknown function, while others were predicted to be regulatory factors and enzymes involved in signaling. We found that bromoform content obtained from female samples (8.5 ± 1.0 mg·g⁻¹ dry weight) was ~10% higher on average than that of male samples (6.5 ± 1.0 mg·g⁻¹ dry weight), although no significant difference was observed (p > 0.05). There was also no significant difference in the marine bromoform biosynthesis locus gene expression. In summary, our comparative RNA-sequencing analysis provides a first insight into the potential molecular factors relevant to gametogenesis and sexual differentiation in A. taxiformis, with potential benefits for identification of sex-specific markers.