性别决定基因SRY的研究进展

SRY基因是哺乳动物性别决定过程中的主宰基因,其表达产物SRY蛋白是一种DNA结合蛋白,该蛋白含有一个HMG盒,能够以序列特异性结合到DNA双螺旋链的一侧,起到转录因子的作用。调节或协同下游基因如SOX9、AMH等基因的表达,使胚胎发育向雄性方向发展。     

Evolution of nodal and nodal‐related genes and the putative composition of the heterodimers that trigger the nodal pathway in vertebrates

Nodal is a signaling molecule that belongs to the transforming growth factor‐β superfamily that plays key roles during the early stages of development of animals. In vertebrates Nodal forms an heterodimer with a GDF1/3 protein to activate the Nodal pathway. Vertebrates have a paralog of nodal in their genomes labeled Nodal‐related, but the evolutionary history of these genes is a matter of debate, mainly because of the presence of a variable numbers of genes in the vertebrate genomes sequenced so far. Thus, the goal of this study was to investigate the evolutionary history of the Nodal and Nodal‐related genes with an emphasis in tracking changes in the number of genes among vertebrates. Our results show the presence of two gene lineages (Nodal and Nodal‐related) that can be traced back to the ancestor of jawed vertebrates. These lineages have undergone processes of differential retention and lineage‐specific expansions. Our results imply that Nodal and Nodal‐related duplicated at the latest in the ancestor of gnathostomes, and they still retain a significant level of functional redundancy. By comparing the evolution of the Nodal/Nodal‐related with GDF1/3 gene family, it is possible to infer that there are several types of heterodimers that can trigger the Nodal pathway among vertebrates.     

Origin of the murine implantation serine proteinase subfamily

The S1 serine protease family is one of the largest gene families known. Within this family there are several subfamilies that have been grouped together as a result of sequence comparisons and substrate identification. The grouping of related genes allows for the speculation of function for newly found members by comparison and for novel subfamilies by contrast. Analysis of the evolutionary patterns of genes indicates whether or not orthologs are likely to be identified in other species as well as potentially indicating that hypothesized orthologs are in fact not. Looking at subtle differences between subfamily members can reveal intricacies about function and expression. Previously, we have described genes encoding two novel serine proteinases, ISP1 and ISP2, which are most closely related to tryptases. The ISP1 gene encodes the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching and invasion in vitro. Additionally both ISP1 and ISP2 are co-expressed in the endometrial gland during the time of hatching, suggesting that they may also both participate in zona lysis from within the uterine lumen. Here, we demonstrate that the ISPs are tandemly linked within the tryptase cluster on 17A3.3. We suggest that remarkable similarities within the 5'-untranslated and first intron regions of ISP1 and ISP2 may explain their intimate co-regulation in uterus. We also suggest that ISP genes have evolved through gene duplication and that the ISP1 gene has also begun to adopt an additional new function in the murine preimplantation embryo.     

植物抗病分子机制研究进展

植物抗病机制是目前研究的热点。在长期的进化过程中,植物形成了一系列复杂有效的防御机制来抵御、破坏病原物的侵染。植物抗病基因在植物抗性反应中起着重要的作用,植物一旦监测到病原物马上起始防御反应,并伴随着植物体内一系列细胞和生理生化的变化。近年来,基因沉默作为一个重要的细胞内防御外源核酸的机制,越来越受到科学家重视。综述了植物抗病基因和基因沉默机制在植物抗病反应中的重要作用,并对研究植物抗病机制的前景做了展望。     

Milbemycin C5-O-甲基转移酶基因的克隆

目的是克隆milbemycin C5-O-甲基转移酶基因。以阿维菌素（avermectin)产生菌S.avermitilis中C5－O－甲基转移酶基因aveD为探针,利用核酸杂交分析法将aveD的主要同源区定位在（22）g-8-E-10阳性克隆所整合的.3kb SatⅠ-BgⅢ片段上。最后对aveD同源区及附近的核苷酸序列进行了测序分析和相应区域的基因缺失分析。结果表明,该区域存在一个月与aveD同源性较强的基因milD。与aveD功能相似,milD参与milbemycin的生物合成,在milbemycin生物合成中可能催化C5位OH的甲基化反应。     

Harnessing the cellular immune system to the gene-prediction cart

Prediction of genes and verification of their bona fide expression in the cell are major challenges of the post-genomic era. Here, we demonstrate how information from the apparently unrelated field of cellular immunology can be recruited for these challenging tasks. The cellular immune system presents short peptides that are the degradation products of both foreign and self-proteins expressed in the cell. We carried out a comprehensive search comparing these peptides to all accumulated human sequence data. Our findings illustrate how these ‘presented self-peptides’ are informative for the identification of new genes, for hypothetical gene verification, for verifying gene expression at the protein level and for supporting splice junctions.     

与宿主昆虫液化相关的杆状病毒基因及其蛋白

昆虫被杆状病毒感染后会发生液化现象,这有利于病毒向周围环境扩散。目前在杆状病毒苜蓿银纹夜蛾核型多角体病毒NPV和GV中,发现与昆虫宿主液化相关的基因有组织蛋白酶基因V-cath基因和几丁质酶基因。V-cath基因表达产物在苜蓿银纹夜蛾多角体病毒(AcMNPV)中能特异性降解昆虫细胞内的肌动蛋白。几丁质酶不仅参与了虫体体表面几丁质的降解,同时还参与V-CATH蛋白前体的加工过程,起分子伴侣的作用。对家蚕核型多角体病毒(BmNPV)的研究表明其FP25K基因表达产物通过影响组织蛋白酶的释放与分泌而参与虫体液化。简要综述了此3种基因及其表达产物的结构、功能与特性,并讨论了它们在生产上的应用前景。     

基因打靶技术:开启遗传学新纪元

基因打靶技术作为最有效的定向修饰小鼠基因的技术手段在揭示基因的生理功能、研究人类疾病的遗传机制以及寻找新的药物靶标的过程中发挥着重要的作用。近年来, 随着条件基因打靶技术的发展使基因失活可以限制在特定时段特定组织或细胞内。文章将主要介绍基因打靶技术的发展简史、近期进展以及在其他模式动物中的应用。     

Cloning and characterization of the rat TIS11 gene

Pirfenidone (Pf), a new broad-spectrum anti-fibrotic agent, is known to offer protection against lung fibrosis in vivo in laboratory animals, and against mitogenesis and collagen formation by human lung fibroblasts in vitro. Because reactive oxygen species are thought to be involved in these events, we investigated the mechanism(s) by which Pf ameliorates oxidative stress and its effects on NADPH-dependent lipid peroxidation. Pf has been shown to cause inhibit NADPH-dependent lipid peroxidation in sheep liver microsomes in a dose-dependent manner. The concentration of Pf required to cause 50% inhibition of lipid peroxidation was ~ 6 mM. Pf was found to be ineffective as a superoxide radical scavenger. Pf was also ineffective in decomposing H₂O₂ and chelating iron. In deoxyribose degradation assays, Pf was a potent scavenger of hydroxyl radicals with a rate constant of 5.4 × 10⁹ M^-1 sec^-1. EPR spectroscopy in combination with spin trapping techniques, using a Fenton type reaction and DMPO as a spin-trapping agent, Pf scavenged hydroxyl radicals in a dose-dependent manner. The concentration of Pf required to inhibit 50% signal height was ~ 2.5 mM. Because iron was used in the Fenton reaction, the ability of Pf in chelating iron was verified in a fluorescent competitive assay using calcein as the fluorescent probe. Pf up to 10 mM concentration was ineffective in chelating either Fe²⁺ or Fe³⁺ in this system. We propose that Pf exerts its beneficial effects, at least in part, through its ability to scavenge toxic hydroxyl radicals.     

The role of circadian clock genes in the photoperiodic timer of the linden bug Pyrrhocoris apterus during the nymphal stage

Many insects survive seasonal adversities during diapause, a form of programmed developmental and metabolic arrest. Photoperiodically regulated entry into diapause allows multivoltine insect species to optimize the number of generations. The molecular mechanism of the photoperiodic timer is unknown in insects. In the present study, we take advantage of the robust reproductive diapause response in the linden bug Pyrrhocoris apterus and explore the fifth‐instar nymphal stage, which is the most photoperiod‐sensitive stage. The nymphs display daily changes in locomotor activity during short days; this differs from the activity observed during long days. We find evidence of cyclical expression of the circadian clock genes, per and cyc, in nymphal heads; in addition, per expression is also photoperiod‐dependent. The RNA interference‐mediated knockdown of the two circadian clock genes, Clk and cyc, during the nymphal stage results in reproductive arrest in adult females. Furthermore, Clk and cyc knockdown induces the expression of the storage protein hexamerin in the fat body, whereas the expression of vitellogenin diminishes. Taken together, these data support the involvement of circadian clock genes in photoperiodic timer and/or diapause induction.     

基因治疗研究中脂质体介导的基因转移技术

对于脂质体的深入研究特别是阳离子脂质体的研制使其逐步成为重要的基因转移载体之一,并且初步应用于基因治疗研究,同时多种靶向脂质体的研制也为体内靶向基因转移和表达奠定了基础。本文就脂质体的结构、功能、在基因治疗研究中的应用以及各种靶向脂质体的研制进行了介绍。     

NAG11和NAG12基因转染对鼻咽癌细胞生长的影响

为了考察鼻咽癌表达下调／缺失基因NAG11和NAG12对鼻咽癌细胞系HNE1生长的影响,构建了NAG11和NAG12基因真核表达载体pcDNA3.1（＋）／NAG11和pcDNA3.1（＋）／NAG12,采用脂质体转染技术将真核重组质粒和空载体质粒分别导入HNE1细胞,观察转染后HNE1细胞生物学特性的变化,结果显示,NAG11重表达对HNE1细胞生长和细胞周期没有明显的影响,而NAG12重表达对HNE1细胞有生长抑制作用,与空载体转染组相比,倍增时间由24.1h延长至31.1h,停滞于G0－G1期细胞数由51.42%增加至68.14%。以上实验进一步说明鼻咽癌是多基因改变的疾病,NAG12的重表达有助于处国咽癌恶性表型的逆转。     

人类基因组中巢式基因对的系统分析

基因组上一个基因的所有或大部分外显子位于另一基因内含子和UTR中被称为巢式基因(Nested gene)。巢式基因对(Nested gene pair)是由主基因(Host gene)和巢式基因组成的基因对。文章针对人类基因组上的顺式巢式基因对(简称为巢式基因对)进行全基因组鉴定,并通过比较巢式基因对在人和小鼠基因组上的保守性分布,研究巢式基因对的进化方式。保守性分析表明基因转座、序列原位突变和主基因转录起始/终止位点突变是巢式基因对进化的主要原因,其中主基因转录起始/终止位点突变是巢式基因对独特的一种进化方式。Gene On-tology分析揭示大部分巢式基因与主基因的产物在功能上没有相关性。     

Evaluation of different target genes for the detection of Salmonella sp. by loop-mediated isothermal amplification

The loop-mediated isothermal amplification (LAMP) technique was used to investigate six salmonella-specific sequences for their suitability to serve as targets for the pathogen identification. Sequences selected for designing LAMP primers were genes invA, bcfD, phoP, siiA, gene62181533 and a region within the ttrRSBCA locus. Primers including single nucleotide polymorphisms were configured as degenerate primers. Specificity of the designed primer sets was determined by means of 46 salmonella and 32 other food- and waterborne bacterial reference species and strains. Primers targeting the ttrRSBCA locus showed 100 % inclusivity of target and exclusivity of other test species and strains. Other primer sets revealed deficiencies, especially regarding Salmonella enterica subsp. II–IV and Salmonella bongori. Additionally, primers targeting the siiA gene failed to detect S. enterica subsp. enterica serotypes Newport and Stanley, whereas bcfD primers did not amplify DNA of S. enterica subsp. enterica serotype Schleissheim. TtrRSBCA primers, providing short detection times and constant melting temperatures of amplification products, achieved best overall performance.     

脊椎动物ABCA基因亚家族研究进展

ABC（ATP-binding cassette）基因家族编码膜蛋白,其成员负责多种物质的跨膜运输。基于氨基酸序列的同源性,人的48个ABC成员被分为7个亚家族：ABCA~ABCG。与其他亚家族相比,ABCA基因编码的蛋白具有独特的拓扑结构,并且其家族成员在两栖动物和哺乳动物分化之后各发生过一次大的扩展（expanding）。基因结构分析发现这两次扩展均是通过基因倍增实现的,这些倍增的产物在啮齿目和食肉目中得到保留,而在灵长目中却有一半变成假基因或被删除。ABCA成员主要负责不同组织器官脂类和胆固醇的跨膜运输,部分成员的突变与疾病相关。     

ht-Pam基因在山羊β-酪蛋白基因座定位整合的研究

利用体细胞基因打靶与核移植技术制备动物乳腺生物反应器是当今转基因定位整合表达的一种新技术。分别克隆山羊的β-酪蛋白基因5′调控区的6.3kb片段,外显子7、外显子8和9三个基因片段,并与克隆的人tPA突变体cDNA一起构建了含有neo和tk正负筛选标记基因的β-酪蛋白基因打靶载体P^GBC4tPA,并验证了neo基因、tk基因以及Cre-LoxP系统的有效性。将线性化的P^GBC4tPA通过电转染整合到山羊胎儿成纤维细胞基因组中,利用G418和GANC进行抗性细胞克隆的药物筛选,初步获得抗性细胞克隆244个,PCR检测后获得阳性细胞克隆31个,其中初步验证2个细胞克隆转植基因整合位点重组后的基因序列正确,并且该细胞克隆能够有效扩增。这为下一步基因打靶体细胞核移植制备山羊乳腺生物反应器奠定了基础。     

植物ASR基因研究进展

ASR（abscisic acid,stress,ripening-induced）基因是近年来从植物中发现的一类受ABA、胁迫和成熟诱导表达的基因,具有保守的ABA/WDS结构域。ASR基因不仅参与植物对干旱、高盐、低温以及脱落酸的胁迫应答,而且参与植物生命活动的许多过程,如果实发育、成熟和糖代谢等。本文综述了近年来国内外ASR基因的研究进展,主要包括ASR基因和蛋白结构特点、ASR基因家族的进化、ASR基因的表达及可能具有的功能,为植物ASR基因研究提供参考。