能在肝癌细胞特异表达的自主复制型肝导向基因转移系统

构建了以人甲胎蛋白基因上游调控序列控制外源基因的表达的自主复制型EB病毒复制子载体pEBAF.该载体与半乳糖基化组蛋白结合组成了一种能为肝细胞表面特异受体识别并内在化的核酸蛋白复合物,能通过肝细胞受体介导的内吞作用以非病毒感染的形式,将外源基因导入细胞内自主复制并仅在产甲胎蛋白的肝癌细胞特异表达.这种新型的基因转移系统,具有比直接使用重组病毒颗粒感染细胞更高的安全性,有应用于原发性肝癌的基因治疗的前景     

Carboxyl terminal of rhodopsin kinase is required for the phosphorylation of photo—activated rhodopsin

Human rhodopsin kinase (RK) and a carboxyl terminus-truncated mutant RK lacking the last 59 amino acids (RKC) were expressed in human embryonic kidney 293 cells to investigate the role of the carboxyl terminus of RK in recognition and phosphorylation of rhodopsin.RKC,like the wild-type RK,was detected in both plasma membranes and cytosolic fractions.The Cterminal truncated rhodopsin kinase was unable to phosphorylate photo-activated rhodopsin,but possesses kinase activity similar to the wild-type RK in phosphorylation of small peptide substrate.It suggests that the truncation did not disturb the gross structures of RK catalytic domain.Our results also show that RKC failed to translocate to photo-activated rod out segments.Taken together,our study demonstrate the carboxyl terminus of RK is required for phosphorylation of photo-activated rhodopsin and strongly indicate that carboxyl-terminus of RK may be involved in interaction with photo-activated rhodopsin.     

丙型肝炎病毒核心蛋白抑制乙型肝炎病毒转录的分子机理

为研究丙型肝炎病毒 (HCV)核心蛋白抑制乙型肝炎病毒 (HBV)表达的分子机理 ,从HCV和HBV共感染中国病人血清中分离了 5个含HCV 5′非编码区 ( 5′NCR)和核心区 (CR)cDNA序列的克隆 .序列比较和系统发育分析显示 :从共感染病例中分离的HCV序列与其它已发表的从单独HCV感染病例中分离的序列没有显著差异 .共转染实验证实采用从其中 1例共感染病人中分离的核心蛋白cDNA基因所表达的核心蛋白 ,可抑制HBV表面抗原 (HBsAg)和HBVe抗原 (HBeAg)的表达 ,缺失突变分析说明HCV核心蛋白的C端疏水区对这种抑制作用是必需的 ,报道基因产物分析进一步说明了HBVC启动子和增强子Ⅱ是HCV核心蛋白的效应靶序列之一 ,而HCV核心蛋白在肝细胞和非肝细胞中均对HBV和其他细胞和病毒的基因的转录起负调节作用 .因此 ,认为HCV核心蛋白是一种多功能的负调节因子 ,与HCV和HBV以及HCV与细胞之间的相互作用密切相关     

Induction of apoptosis and change of bcl—2 expression in macrophage Ana—1 cells by all—trans retinoic acid

Macrophage cells play an important role in the initiation and regulation of the immune response.All-trans retinoic acid (ATRA) and its natural and synthetic analogs (retinoids)affect a large number of biological processes.Recently,retinoids have been shown promise in the therapy and prevention of various cancers.However,many interesting questions related to the activities of retinoids remain to be answered:(I) Molecular mechanisms by which retinoids exert their effects;（Ⅱ)why the clinical uses of retinoids give undesirable side effects of varying severity with a higher frequency of blood system symptoms;(Ⅲ)little is known for its impacts on macrophage cells etc.We set up this experiment,therefore,to examine the apoptosis of ATRA on macrophage Ana-1 cell line.Apoptosis of the cells was quantitated,after staining cells with propidium iodide(PI),by both accounting nuclear condensation and flow cytometry.When the cells were treated with ATRA at or higher than 1μM for more than 24h,significant amount of the apoptotic cells was observed.Induction of apoptosis of Ana-1 cells by ATRA was in time-and dose-dependent manners,exhibiting the similar pattern as the apoptosis induced by actinomycin D (ACTD).ATRA treatment of Ana-1 cells also caused the changes of the mRNA levels of apoptosis-associated gene bcl-2,as detected by Northern blot analysis.The temporal changes of bcl-2 expression by ATRA was also parallel to that by ACTD.In conclusion,ATRA can induce apoptosis in macrophage cells,which may be helpful in understanding of immunological functions retinoids.     

稻田节肢动物群落多样性

本文运用种类丰富度(S)、均匀性(V)和申农指数(Shannon's index.H')分析稻田节肢动物群落和各亚群落多样性的空间层次差异、季节动态、以及杀虫药剂的作用.稻田节肢动物群落的多样性值较低,H'值一般在3以下,表明群落稳定性差;多样性季节变化较大.在不用农药条件下,8月底以后的H'值较高;水稻群体上部群落的S、和V'值均大于下部的对应值;杀虫药剂的应用虽提高了群落总体的H'和V'值,但对亚群落的分析表明,杀虫药剂促进害虫亚群落的H'和V'值上升,却降低了各天敌亚群落的有关值,从而扩大了害虫和天敌之间稳定性的差距,使两者的同步关系受到损害;杀虫药剂的应用还使各天敌亚群落种类数和个体数明显减少,特别是寄生天敌昆虫种类数、个体数均大幅度下降,使其难以起到控制害虫的应有作用,这又为害虫的再猖獗提供有利条件.通过分析认为,就稻田节肢动物而言,多样性仍然是群落稳定性的一个重要测定指标.     

Embryology of Plagiopteron suaveolens Griffith (Plagiopteraceae) and its systematic implications

A study of the embryology of Plagiopteron suaveolens Griffith is presented in order to evaluate the affinities of the genus. Anther wall formation is of the dicotyledonous type. The anther tapetum is glandular and its cells become two to four nucleate. Cytokinesis in the microspore mother cell is simultaneous and the resultant tetrad is tetrahedral. The mature pollen grains are two-celled. The anther has 4-locules at anthesis. The ovule is anatropous, bitegmic and weakly crassinucellate with only one cell-layered parietal tissue. The micropyle is zig-zag in section and formed by both inner and outer integuments which are three and three to four cell layers thick respectively. The megaspore archesporium is one-celled and the embryo sac development is of the Polygonum type. An endothelium is formed with early disintegration of nucellar tissue. The antipodals multiply to 15 to 20 cells. Embryological evidence supports the placement of Plagiopteron in its own family, Plagiopteraceae, with closest relationship with Celastraceae and close relationship with Elaeocarpaceae, Rhizophoraceae and some families of the Geraniales.     

一个含端粒序列的水稻BAC克隆的分析和定位

对一个含 (TTTAGGG) _n 同源序列的水稻BAC克隆 (BAC2 )进行了分析 ,揭示出水稻近端区DNA的组成 .BAC2的插入片段中除含有大量的以串联形式存在的称之为TA35 2序列的卫星DNA外 ,还含有TTTAGGG或其变体组成的简单重复 .荧光原位杂交 (FISH)将含 (TTTAGGG) _n 序列的 0 .8kbPstⅠ片段定位在至少 5对染色体的端粒区 .通过对BAC2中低拷贝序列的RFLP分析 ,BAC2被定位在水稻第 6号染色体端部 .这些结果说明水稻的端粒序列可能也是TTTAGGG或其变体构成的简单重复 ,而与其相连的卫星DNATA35 2则属于端粒相关序列 .     

微藻光密度与细胞密度及生物质的关系

以四种常见微藻,小球藻(Chlorella sp.XQ-20044)、栅藻(Scenedesmus sp.SS-200716)、绿球藻(Chlorococcum sp.)和螺旋藻(Spirulina sp.CH-164)为实验材料,用梯度稀释法测定对数生长期不同浓度藻液的光密度(OD)、细胞密度和生物质干重(DW),在光自养分批培养模式下对4种微藻进行OD-波长(350—800 nm)扫描,同时测定细胞密度和生物质干重,分析藻液OD与细胞密度、生物质干重的关系。结果表明:在任何波长下,对数生长期的4种微藻细胞密度与OD值、生物质干重与OD值的变化都不成比例,波长不同其拟合曲线偏离直线的程度不同。但是,在435 nm处这种关系最接近直线,可以用直线方程近似描述(R20.98),其它波长处细胞密度-OD、干重-OD的关系都可以用二项式方程很好地描述(R20.99)。因此,光密度法适用于连续和半连续培养,可以用435 nm处测得的OD值计算细胞密度与干重。但是在分批培养模式下,4种微藻DW/OD比值随着培养时间均逐渐上升。小球藻DW/OD540为0.19—0.44 g/L,栅藻DW/OD540为0.36—0.53 g/L,绿球藻DW/OD540为0.48—0.75 g/L,螺旋藻DW/OD560为0.46—0.74 g/L,因此分批培养模式下采用测定藻液OD值反映细胞密度和生物质的方法不适用,只有直接测定细胞密度和生物质才是准确的。研究结果为正确使用分光光度法监测微藻生长提供依据。     

红火蚁三龄、四龄幼虫肠道菌菌群多样性分析

昆虫体内细菌的多样性对昆虫的消化、营养的吸收、生长发育繁殖具有至关重要的作用.本研究利用16S rRNA基因序列,构建内生菌16S rRNA克隆文库,探明红火蚁3龄幼虫和4龄幼虫种群体内微生物菌群结构并加以对比,采用Miseq高通量测序的方法揭示红火蚁3龄幼虫与4龄幼虫肠道菌的相似性和差异性.结果表明:红火蚁高龄幼虫的...     

宏基因组来源耐Mn2+、热稳定细菌漆酶的分子克隆及酶学特性

【背景】漆酶和锰过氧化物酶(Manganese peroxidase,Mnp)是木质素降解的主要酶,二者有协同效应。Mnp活性依赖于Mn~(2+),而Mn~(2+)是大多数漆酶的抑制剂。【目的】获得耐Mn~(2+)的细菌漆酶用于木质素降解。【方法】构建许昌市某污水河污泥宏基因组文库,通过活性筛选获得细菌漆酶基因lac1542。使用大肠杆菌异源表达Lac1542,研究纯化后的重组蛋白酶学性质并进一步检测了含Lac1542复合酶系降解木质素能力。【结果】测序结果显示lac1542编码一个含513个氨基酸的蛋白。以ABTS为底物Lac1542最适反应pH为4.0,在pH 3.0-6.5范围内酶活性稳定。最适反应温度是75°C,在70°C以下酶活性稳定;100 mmol/L的Mn~(2+)仍能提高酶的活性。动力学参数研究发现,该酶的最适底物顺序为:ABTS丁香醛联氮儿茶酚2,6-DMP愈创木酚。Lac1542/Mnp复合酶系对木质素降解率为47.8%,比单独使用Mnp木质素降解率(22.4%)提高25.4%。Lac1542/Mnp/灰盖鬼伞过氧化物酶(Coprinus cinereus Peroxidase,CIP)复合酶系木质素降解高达71.5%,比Mnp/CIP酶系木质素降解率(48.9%)提高22.6%,加入Lac1542后的复合酶系能明显提高木质素的降解率。【结论】Lac1542的可溶性表达、耐受高浓度Mn~(2+)、热稳定性使得Lac1542可以替代一些经典的真菌漆酶应用于制浆、造纸、纤维素乙醇生产、染料脱色等工业。