大鼠尿激酶SYBR Green I real time RT-PCR引物的设计

设计用于SYBR Green I法实时定量逆转录多聚酶链反应(QRT-PCR)检测大鼠尿激酶型纤溶酶原激活因子(uPA)mRNA的引物。从基因库获取靶基因及相关序列,充分收集争分析相关生物信息学数据,应用Oligo 6.22设计出一对长度为21bp的引物,其GG含量为52.4%;上下游引物3’最稳定二聚体和及发夹结构的能量分别为-1.5、-0.40 kcal/mol和-3.5、-O.90 kcal/mol,引物间最稳定二聚体为-3.1 kcal/mol。5’端和中间△G值较高,高于3’端△G;引发效率分别455和403。实验证明,该引物能够高效、特异地实现对靶序列的检测,适用于SYBR Green I法实时定量检测(uPA)mRNA。     

肾上腺素对THP-1巨噬细胞TGF-β1、SR-A1和CD36表达的影响

目的:探讨肾上腺素对THP-1巨噬细胞TGF-β1、SR-A1和CD36表达的影响。方法:不同浓度肾上腺素处理THP-1巨噬细胞24h,运用逆转录多聚酶链反应检测其TGF-β1、SR-A1和CD36的mRNA表达,酶联免疫吸附试验检测TGF-β1蛋白的表达,Westem-blotting检测SR-A1蛋白的表达。结果:100nmol/L及1μmol/L的肾上腺素作用THP-1巨噬细胞,引起TGF-β1 mR- NA和蛋白表达下调(p<0.05),SR-A1 mRNA和蛋白表达上调(P<0.05),1pmol/L～1μmol/L的肾上腺素处理后,各组CD36 mRNA水平无显著性变化(p>0.05)。结论:应激浓度的肾上腺素可能通过影响TGF-β1和SR-A1的表达而参与和/或促进AS的发生发展。     

太子参花药发育及精细胞分离

太子参花药壁发育为基本型,腺质绒毡层。小孢子母细胞减数分裂为同时型,小孢子四分体为四面体型,成熟花粉具两个精细胞,为3胞花粉。在花粉表面具散孔,孔数22—30个,均匀分布于花粉粒表面上。花粉在10％甘露醇或15％蔗糖溶液中可直接爆破,精细胞易被释放并散开,通过显微操作仪可收集到一定数目的精细胞。FDA染色荧光显示释放出来的精细胞活力可维持25—50min。花粉在舍O．03％CaCl2、0．01％H3803、0．01％KH2P04和20％PEG、pH5．8的培养液中2—5min即萌发花粉管．花粉管生长2h可达815μm。一般花粉管伸长500—600μm时,一对精细胞才进入花粉管。DAPI染色后荧光观察．可观察到精细胞和营养细胞核在花粉管中的移动状况。爆破花粉管后可释放出一对精细胞。     

紫茎泽兰入侵对土壤酶活性和理化因子的影响

紫茎泽兰是我国危害最严重的外来入侵植物之一,为探明其入侵对土壤肥力的影响,比较研究了紫茎泽兰、云南菅、狗尾草群落和撂荒地下0~30 cm的4层土壤中6种酶活性和12种理化因子。结果表明群落类型和土壤深度对测定的各参数均有显著影响。随土壤深度的增加,多酚氧化酶、碱性磷酸酶、脲酶活性,以及有机质、全氮、全磷、全钙、水解氮、有效磷、速效钾含量和pH值均降低。总的看来,紫茎泽兰群落下碱性磷酸酶和脲酶活性,有机质、全氮、全磷、全钙、水解氮和有效磷含量,以及pH值均较高,全钾含量较低,但速效钾含量并不低,表明紫茎泽兰入侵多年后土壤肥力水平提高,形成了对其生长有利的土壤环境。     

人血清蛋白质组学方法的比较和优化

目的:对人未做处理的血清以及去除白蛋白和免疫球蛋白G(IgG)血清的蛋白质组学方法进行比较和优化。方法:应用双向电泳(2-DE)方法分离了未做处理的以及去除白蛋白和免疫球蛋白G(IgG)的血清,比较优化了高温变性、水化液成份组成及泡胀方式等影响血清2-DE分离效果的因素,并用质谱分析鉴定未做处理和已处理血清的2-DE谱图中部分差异蛋白点。结果:得到了分辨率和重复性较好的2-DE谱图,未做处理的血清、去除白蛋白及IgG血清的平均蛋白质点分别为(482±18)个和(523±29)个,质谱分析了9个差异蛋白点,鉴定为8种蛋白质,其中7种为功能蛋白质。仅出现在未做处理血清中的蛋白有4种,分别是维生素A结合蛋白、可溶性尿激酶血纤维蛋白溶酶原激活剂受体、蛋白激酶1抗原、血清白蛋白。4种蛋白仅出现在去除白蛋白和IgG的血清中,分别是NADH脱氢酶辅酶β亚基、肌动蛋白结合蛋白M1、T细胞活性受体β链、血小板生长因子C。结论:去除高丰度蛋白可增加一些低丰度蛋白质的检出,但非特异性吸附会导致部分功能蛋白质的丢失。     

注射性组织工程牙胚的构建及成牙能力研究

目的:探索采用大鼠牙胚细胞构建可注射组织工程牙齿的可行性及在体内肾被膜移植的成牙能力研究。方法:采用出生后4d SD仔鼠磨牙牙胚细胞,培养传代后将牙胚细胞与牛真皮基质蛋白溶液混合构建组织工程牙胚,利用注射器植入SD大鼠肾被膜下,2wk后取材,HE染色观察移植物中的组织成熟情况,Masson’s三色法观察组织工程牙胚的成牙能力。结果:组织工程牙胚移植后2wk可在肾被膜下形成乳白色矿化组织,HE染色上皮细胞和间充质细胞进行重组,形成典型的牙髓牙本质复合体样组织和釉质样结构,Masson’s三色法可见绿色矿化基质形成并包绕牙乳头样组织。结论:牙胚细胞仍保留了牙齿发育的位置信息,采用牛真皮基质蛋白溶液作为组织工程支架有助于牙胚细胞相互作用并重新排列,构建的组织工程牙胚具有进一步成熟、矿化并形成牙齿的潜能。     

洋葱花粉发育过程中ATP酶的分布特征(简报)

在真核细胞中,除了线粒体和叶绿体ATPase的功能是合成ATP外,其余部位ATPase是水解ATP以获取生物能量的代谢酶,在生物体细胞内广泛存在。探索ATPase在细胞中的分布状态是研究细胞生理状态的一种重要手段。ATPase在细胞中的多少可反映出细胞当时的生活状态,这一特征已被初步用于探索小麦和水稻雄性不育的细胞生物学研究中,希望通过比较可育花药和不育花药中ATPase的分布差异寻找雄性不育的机理,发现     

一个Ⅰ型遗传性淋巴水肿中国家系的VEGFR3基因的新突变方式

通过对国人Ⅰ型遗传性淋巴水肿一家系分子遗传学检测,报告VEGFR-3基因新突变。首先在Ⅰ型遗传性淋巴水肿对该家系进行致病基因的连锁分析,然后用DNA直接测序方法进行基因突变分析。连锁分析和单倍体分析确定该家系致病基因位于5q35.3,与Ⅰ型遗传性淋巴水肿连锁。VEGFR-3基因突变分析发现了一个新的错义突变D1055V,该错义突变在家系中共分离,且在100个正常对照组中未发现该序列改变。本研究首次报告了国内Ⅰ型遗传性淋巴水肿VEGFR-3基因新的错义突变D1055V,丰富了VEGFR-3基因基因突变谱,为今后开展遗传性淋巴水肿的基因诊断和遗传咨询奠定基础。     

Changes in biodiversity and ecosystem function during the restoration of a tropical forest in south China

Tropical forests continue to vanish rapidly,but few long-term studies have ever examined if and how the lost forests can be restored.Based on a 45-year restoration study in south China,we found that a tropical rain forest,once completely destroyed,could not recover naturally without deliberate restoration efforts.We identified two kinds of thresholds that must be overcome with human ameliorative measures before the ecosystem was able to recover.The first threshold was imposed primarily by extreme physical conditions such as exceedingly high surface temperature and impoverished soil,while the second was characterized by a critical level of biodiversity and a landscape context that accommodates dispersal and colonization processes.Our three treatment catchments(un-restored barren land,single-species plantation,and mixed-forest stand)exhibited dramatically different changes in biodiversity and ecosystem functioning over 4 decades.The mixed forest,having the highest level of biodiversity and ecosystem functioning,possesses several major properties of tropical rain forest.These findings may have important implications for the restoration of many severely degraded or lost tropical forest ecosystems.     

Comparison between gene expression of conidia and germinating phase in Trichophyton rubrum

Trichophyton rubrum is a dominating superficial dermatophyte, whose conidial germination is correlated to pathopoiesis and a highly important developmental process. To investigate the changes of physiology, biochemistry and cytology during the germination, we selected 3364 function identified ESTs from T. rubrum cDNA library to construct cDNA microarrays, and compared the gene expression levels of conidia and germinating phase. Data analysis indicated that 335 genes were up-regulated during the germination, which mainly encoded translated, modified proteins and structural proteins. The constituents of cell wall and cell membrane were synthetized abundantly, suggesting that they are the foundation of cell morphogenesis. The ingredients of the two-component signal transduction system were up-regulated, presuming that they were important for the conidial germination. Genes of various metabolic pathways were expressed prosperously, especially the genes that participated in glycolysis and oxidative phosphorylation were up-regulated on the whole, demonstrating that in the environment with sufficient oxygen and glucose, conidia obtained energy through aerobic respiration. This paper provides important clues which are helpful to understanding the changes in gene expression, signal conduction and metabolism characteristics during T. rubrum conidial germination, and possess significant meaning to the study of other superficial dermatophytes. Supported by the National High Technology Research and Development Program of China (Grant No. 2001AA223021) and National Key Technologies R&D Programme (Grant No. 2002BA711A14)