重组肾上腺髓质素表达载体的构建及在哺乳动物细胞中的表达

为探索通过体内表达肾上腺髓质素（adrenomedullin,AM）治疗高血压和慢性心衰的可能性,本实验构建了重组AM真核表达载体,并在无内源笥AM表达的K562细胞株上进行了体外表达实验。实验中采用RT－PCR技术扩增AM cDNA片段,并将扩增的cDNA片段插入pcDNA3.1真核表达质粒,构建成含AM cDNA的重组质料pcDNA3.1AM。用脂质体介导将该质粒转染培养的人白血病细腻K562株,在转染的细胞中,用RT－PCR检测证实有AM mRNA存在;用班点免疫分析方法检测转染细胞的培养液上清,证实有AM多肽存在,表明本实验中构建的重组pcDNA3.1AM载体能够在哺乳类细胞中表达AM。     

Changes in Florets’ Vertical Direction within Inflorescence Affects Pollinator Behavior,and Fitness in <i>Trifolium repens</i>

Ecological interactions between flowers and pollinators greatly affect the reproductive success. To facilitate these interactions, many flowers are known to display their attractive qualities, such as scent emission, flower rewards and floral vertical direction, in a rhythmic fashion. However, less is known about how plants regulate the relationship between these flower traits to adapt to pollinator visiting behavior and increase reproduction success. Here we investigated the adaptive significance of the flower bending from erect to downward in Trifolium repens. We observed the flowering dynamic characteristics (changes of vertical direction of florets, flowering number, pollen grain numbers, pollen viability and stigma receptivity over time after blossom) and the factors affecting the rate of flower bending in T. repens. Then we altered the vertical direction of florets in inflorescence of different types (upright and downward), and compared the pollinator behaviors and female reproductive success. Our results showed that florets opened sequentially in inflorescence, and then bend downwards slowly after flowering. The bending speed of florets was mainly influenced by pollination, and bending angle increased with the prolongation of flowering time, while the pollen germination rate, stigma receptivity and nectar secretion has a rhythm of “low-high-low” during the whole period with the time going. The visiting frequency of all the four species of pollinators on upward flowers was significantly higher than that of downward flowers, and they especially prefer to visit flowers with a bending angle of 30°–60°, when the flowers was exactly of the highest flower rewards (nectar secretion and number of pollen grains), stigma receptivity and pollen germination rate. The seed set ratio and fruit set ratio of upward flowers were significantly higher than downward flowers, but significantly lower than unmanipulated flowers. Our results indicated that the T. repens could increase female and male fitness by accurate pollination. The most suitable flower angle saves pollinators’ visiting energy and enables them to obtain the highest nectar rewards. This coordination between plants and pollinators maximizes the interests of them, which is a crucial factor in initiating specialized plant-pollinator relationships.     

Differential ConA-enriched urinary proteome in rat experimental glomerular diseases

Glomerular diseases are leading causes of end-stage renal diseases worldwide. They are considered to be consequences of injury primarily to the three types of glomerular cells. Differential diagnosis typically relies on invasive biopsy findings. We expected that injuries of different glomerular cells would cause different changes in urinary proteome. The goal of this study was to identify differential urinary proteins distinguishing between injuries of different glomerular cells before significant histopathologic changes. Adriamycin nephropathy and Thy1.1 glomerulonephritis were employed as models with different primary impaired cells. ConA-enriched urinary glycoproteome on day3 were profiled by gel-free shotgun tandem mass spectrometry, and compared with self-healthy controls to identify differential urinary proteins for each model. By comparing the changes of the differential proteins between these two models, we identified 39 proteins with different directions of changes, which may potentially be useful in differentiation; and 7 proteins with the same direction of changes, which may be potential indicators of early renal damage. These differential proteins were of several origins: plasma proteins, proteins with urine or kidney specificity, proteins without tissue-specificity (mainly inflammatory mediators) etc. Our results may help better understand the effects of injuries of different glomerular cells at the initial stage, and lead to the discovery of novel early diagnostic markers for human focal segmental glomerulosclerosis (FSGS) and mesangioproliferative glomerulonephritis (MsPGN) which have the same primary impaired cells with adriamycin nephropathy and Thy1.1 glomerulonephritis, respectively.     

Develope Monoclonal Antibody against Foot-and-mouth Disease Virus A Type

In order to develop an anti-FMDV A Type monoclonal antibody (mAb),BABL/c mice were immunized with FMDV A type.Monoclonal antibodies (mAbs) 7B11 and 8H4 against Foot-and-mouth disease virus (FMDV) serotype A were produced by fusing SP2/0 myeloma cells with splenocyte from the mouse immunized with A/AV88.The microneutralization titer of the mAbs 7B11 and 8H4 were 1024 and 512,respectively.Both mAbs contain kappa light chains,the mAbs were IgG1.In order to define the mAbs binding epitopes,the reactivity of the...     

蜜蜂蜂王与工蜂级型分化研究进展

蜜蜂ApismekiferaL .是典型的社会性昆虫 ,蜂王和工蜂都是由受精卵发育而来的二倍体成蜂 ,但是在形态、生理、行为等方面有明显的差异 ,属于不同的级型。蜂王和工蜂的级型分化的关键时期发生在幼虫的 4龄末至 5龄止。分化是由分化基因调控的 ,幼虫期食物的质和量是分化的外部决定因子。JH对两级型中卵巢的分化有非常重要的调控作用。蜜蜂脑或其它组织中可能有分泌调控CA的咽侧体调节激素 ,它们通过对CA中JH的合成和分泌的调控而参与了分化的调控。章鱼胺等生物胺也参与了分化调控过程。     

Human DNA ligases I and III, but not ligase IV, are required for microhomology-mediated end joining of DNA double-strand breaks

DNA nonhomologous end-joining (NHEJ) and homologous recombination are two distinct pathways of DNA double-strand break repair in mammalian cells. Biochemical and genetic studies showed that DNA ends can also be joined via microhomology-mediated end joining (MHEJ), especially when proteins responsible for NHEJ, such as Ku, are reduced or absent. While it has been known that Ku-dependent NHEJ requires DNA ligase IV, it is unclear which DNA ligase(s) is required for Ku-independent MHEJ. In this study, we used a cell-free assay to determine the roles of DNA ligases I, III and IV in MHEJ and NHEJ. We found that siRNA mediated down-regulation of DNA ligase I or ligase III in human HTD114 cells led to impaired end joining that was mediated by 2-, 3- or 10-bp microhomology. In addition, nuclear extract from human fibroblasts harboring a mutation in DNA ligase I displayed reduced MHEJ activity. Furthermore, treatment of HTD114 nuclear extracts with an antibody against DNA ligase I or III also significantly reduced MHEJ. These data indicate that DNA ligases I and III are required in MHEJ. DNA ligase IV, on the contrary, is not required in MHEJ but facilitates Ku-dependent NHEJ. Therefore, MHEJ and NHEJ require different DNA ligases.     

四氯化碳损伤成年大鼠肝细胞后原癌基因（c－fos／c－jun）表达的观察

用四氯化碳（ＣＣｌ４）损伤正常大鼠后,采用Ｗｅｓｔｅｒｎ印迹法和免疫组化法观察肝细胞原癌基因（ｃ－ｆｏｓ／ｃ－ｊｕｎ）的表达。Ｗｅｓｔｅｒｎ印迹法表明,当成年大鼠的静息期肝细胞受到ＣＣｌ４损伤性刺激后,ｃ－ｆｏｓ／ｃ－ｊｕｎ产物（Ｆｏｓ和Ｊｕｎ）水平升高,在ＣＣｌ４处理后３０ｍｉｎ开始升高,在４ｈ时消失。８ｈ后Ｆｏｓ／Ｊｕｎ再度出现,并持续２４ｈ以上。ＩＣＣ法表明,Ｊｕｎ阳性细胞为靠近肝中央静脉区的肝实质细胞。根据上述资料推测,肝受ＣＣｌ４损伤后肝细胞的原癌基因ｃ－ｆｏｓ／ｃ－ｊｕｎ出现即时的与滞后的两次表达,这与肝细胞进入细胞周期有关,这种基因表达也许可作为肝再生过程中识别特殊体液因子的标志。     

益生菌在结直肠癌患者中的应用

结直肠癌(colorectal cancer,CRC)的发病率及病死率在多国多年居高不下,肠道微生态的失衡在CRC的发生发展中所起的作用被许多学者所证实。专家一致认为,积极纠正肠道微生态失衡是可取的。CRC患者术前肠道菌群已经出现改变,术后肠道菌群失衡加重,化疗会进一步加重这种失衡状态。肠道微生态的稳态对机体肠道功能和免疫功能等起着重要作用。益生菌作为一种可调节肠道菌群的微生态制剂,已显现出在CRC患者治疗中的应用价值,现对益生菌在CRC患者中的应用进展作一综述。     

子宫颈糜烂病毒病因的探讨

491份宫颈拭子病毒分离结果表明:糜烂宫颈单纯疱疹病毒(HSV)分离阳性率(30.8％)是正常宫颈(2.6％)的11.8倍,用人干扰素治疗一个疗程后,病毒分离率下降至疗前的1/4.36例糜烂宫颈活体组织DNA分子杂交表明,乳头瘤病毒16型(HPV-16)阳性者占52.8和,HPV-18占17.9％,HPV-6B占28.1％,HPV-11占7.7％,251例宫颈糜烂患者经人(?)D型基因工程干扰素双盲对比治疗后,总有效率达93.8％,显效率达60％,分析临床疗效与HSV分离率的变化表明,临床有效病例中有35％(49/140)在治疗后病毒阴转,有57％在疗前疗后均未分离出HSV,有5％在疗前疗后保持阳性不变,有2.9％疗前阴性,疗后阳性,上述结果表明,HSV和HPV与慢性宫颈炎有一定关系。     

SYBR Green I荧光RT-PCR检测犬瘟热方法的建立和应用

根据GenBank发表的犬瘟热病毒（CDV）的F基因序列,经过分析在F片段的保守区域内设计引物,建立了SYBR Green I荧光RT-PCR检测CDV的方法,并通过对厦门市宠物医院收集的临床发病和疑似发病的犬病料（包括眼分泌物、鼻拭子、唾液、血液、尿液等）的检测,结果表明,本研究建立的快速检测CDV的SYBR Green I荧光RT-PCR方法具有特异性强、灵敏度高、操作简便等优点,值得推广应用。