噬菌体PaP2推定基因功能的研究

一个生物体基因组测序工作的完成 ,是进行基因组学和功能基因组学研究的基础 ,只是巨量工作的开始。本文通过运用GeneMark以及softberryfgenesV等软件在铜绿假单胞菌噬菌体PaP2基因组上共发现了 5 8个推定基因的编码区 ,为了解这些基因的功能 ,进行了两个方面的研究。一方面是将纯化噬菌体颗粒进行SDS PAGE分析 ,发现噬菌体PaP2至少含有 10个衣壳蛋白质 ,再转印到PVDF膜上 ,分别剪下各条带进行蛋白质N 末端氨基酸测序。根据蛋白质实测分子量 ,从 5 8个ORFs中找出编码相近分子量产物的ORFs ,再根据N 端氨基酸序列确定衣壳蛋白质的…     

噬菌体PaP3末端酶大亚单位的克隆表达

末端酶 (terminase)是双链DNA病毒包装过程中 ,将病毒基因组多串联体 (concatemer)与病毒前衣壳连接 ,切割并转移单拷贝基因组进行前衣壳 (procapsid)的异源寡聚体 ,由一个大亚单位及 1～ 7个小亚单位组成。大亚单位具有ATP酶和核酸酶活性 ,在全酶活性中起到重要的作用。由我室分离并完成测序的绿脓杆菌噬菌体PaP3(GeneBank序列号 :AY0 78382 )为双链DNA病毒 ,其中4 0 6 93 4 2 14 1基因编码末端酶大亚单位。本研究采用PCR技术从噬菌体PaP3基因组中扩增出编码噬菌体PaP3末端酶大亚单位的目标片段 ,将目标片段插入原核表达载体 pQE…     

禾谷缢管蚜体内的病毒结合蛋白基因的克隆与原核表达

利用一对特异性引物,用PCR的方法从禾谷缢管蚜体内扩增出了病毒结合蛋白基因,序列测定结果表明其长度 为1647 bp,编码548个氨基酸,与GenBank中的禾谷缢管蚜美国生物型Buchnera groELNT核苷酸序列同源性为97%,氨基酸同源性为97.4%。构建了2个原核表达载体并进行表达得到了69kD融合蛋白和63kD的非融合蛋白。     

MS2噬菌体衣壳蛋白与包装位点结合特异性及其生物学应用进展

MS2噬菌体为正义单链RNA噬菌体,基因组含有3569个核苷酸,编码成熟酶蛋白、衣壳蛋白、复制酶蛋白和裂解蛋白。MS2噬菌体复制酶编码基因5'端一个由19个碱基组成的茎环结构(又称包装位点)是衣壳蛋白二聚体与RNA相互作用的部位,二者相互作用形成的复合物是启动噬菌体自我包装的信号。MS2噬菌体衣壳蛋白与包装位点结合的特异性已被应用于RNA病毒核酸检测的标准物质、校准品和质控品的研究,实时动态监测活细胞内RNA的运动,以及RNA体内递送载体的研究等领域。     

我国蚊虫分离的一种新型胞质多形体病毒

0507BS3是从中国新疆喀什地区采集的库蚊和按蚊混合蚊标本分离的病毒株,对C6/36细胞致病变而对Ve-ro和BHK-21细胞不致病变。电镜观察显示病毒颗粒呈圆球形,直径约60nm(n=20),无包膜,单层衣壳,衣壳内有中央核。基因组核酸电泳显示基因组包括10条双链RNA(double stranded RNA,dsRNA)片段。病毒第10基因片段核酸序列测定显示该片段全长964bp(GenBankID:FJ150869),具有单一开放读码框,编码长度为275个氨基酸的蛋白,分子量约30.8kD。病毒第10基因片段核酸序列比对未发现相似的病毒核酸序列,氨基酸序列与胞质多形体病毒(Cytoplasmic polyhedrosis virus,CPV)第10基因片段编码的多形体蛋白部分区段匹配。病毒第10基因片段和已知各型CPV第10基因片段核酸序列共同进行系统进化分析显示该病毒位于独立的进化分枝,提示0507BS3病毒可能是一种新型CPV病毒。     

从云南烟草上检测到的黄瓜花叶病毒亚组II分离物

在对云南省烟草病毒病的研究中,分离到一种直径约26～30nm的球形病毒。提纯病毒进行的SDSPAGE发现一条55kD蛋白带。55kD蛋白N端10个氨基酸与CMV亚组II的Q株系外壳蛋白N端氨基酸同源性为100%。以CMVQ抗血清对55kD蛋白进行了Western blot检测,发现55kD蛋白与CMV Q株系抗血清有血清学反应。根据已报道的CMV亚组II外壳蛋白基因序列合成引物,采用RTPCR技术扩增到一条约09kb的cDNA条带,并进行了克隆及序列测定,经Genbank比较,发现此09kb cDNA包含一657bp的外壳蛋白基因,其核苷酸序列及其推导的氨基酸序列与CMV亚组II分离物有极高的同源性,分别达97%～98%和96%～99%,而与亚组I分离物的同源性仅为75%～76%和78%～79%。因此,该球形病毒应为CMV亚组II的一个分离物,命名为CMVYnb。     

巴西橡胶树43 kD橡胶粒子膜蛋白基因的cDNA克隆及表达

对43 kD的橡胶粒子膜蛋白进行了分离纯化和其N端氨基酸序列分析,根据N端氨基酸序列,设计一简并引物,通过3'RACE(Rapid Amplification ofcDNA Ends)的方法,获得了43 kD的橡胶粒子膜蛋白的cDNA.该cDNA含有1 385个核苷酸,含有完整的阅读框架,编码381个氨基酸.在终止密码子下游,包含有一个239bp的3'非编码区.该cDNA由5个首尾相连的重复单元组成,每个单元编码76个氨基酸组成的泛素(ubiquitin)单体.编码43 kD橡胶粒子蛋白的基因具有多个拷贝,在胶乳、叶片和树皮都表达.     

柠条锦鸡儿CkLEA4基因克隆及表达分析

LEA蛋白是一种与植物抗逆相关的胚胎发育晚期丰富蛋白。该研究从已构建的柠条锦鸡儿干旱胁迫抑制削减杂交文库中筛选到1条LEA蛋白编码基因的部分序列,用RACE技术扩增得到该基因cDNA全长并对其进行了克隆。测序表明该基因cDNA长870bp,其中开放阅读框长510bp,编码169个氨基酸,推测蛋白分子量为17.03kD,等电点为9.3,是一种亲水蛋白。序列比对和系统进化分析表明,该基因属于LEA4基因家族成员,命名为CkLEA4。实时荧光定量PCR检测发现,CkLEA4基因在干旱、ABA和NaCl处理下均受到不同程度的诱导,说明CkLEA4基因可能与柠条锦鸡儿响应逆境胁迫有关。     

柞蚕14-3-3基因的鉴定及表达分析

14-3-3蛋白是一种可以改变其结合蛋白构象的酸性蛋白质.柞蚕14-3-3 cDNA序列全长1 220 bp,包括一个126 bp的5'非编码区和一个350 bp的3'非编码区.该基因的开放读码框长度为744 bp,编码247个氨基酸.序列比对结果表明,柞蚕14-3-3蛋白与家蚕的14-3-3蛋白具有高度同源性.此外对柞蚕14-3-3基因进行了原核表达和重组蛋白纯化.SDS-PAGE和免疫印迹结果表明,分子量大小约32 kD的重组蛋白在大肠杆菌中得到了成功表达.     

传染性支气管炎病毒核衣壳蛋白基因克隆及在E.coli中的表达

核衣壳蛋白基因 (N基因 )是传染性支气管炎病毒的重要结构基因 .根据已报道的序列设计引物 ,利用RT PCR技术从病毒RNA中扩增和克隆到了N基因的cDNA ,并测定了核苷酸序列 .克隆的N基因片段ORF全长 12 30bp ,编码 4 0 9个氨基酸 .将该片段序列与其他IBV病毒株比较 ,核苷酸的同一性为 87 0 %～ 98 6 %,氨基酸的同一性为 91 0 %～ 98 1%.将该cDNA亚克隆到pBV2 2 0表达载体 ,转化大肠杆菌DH5α菌株 ,Western印迹检测 ,获得了分子量约 4 5kD表达蛋白     

六种藓类植物茎的比较解剖学研究

通过对6种藓类植物,即褶叶青藓(Brachythecium salebrosum(Web.et Mohr.)B.S.G.)、湿地匐灯藓(Plagiomnium acutum(Lindb.)Kop.)、侧枝匐灯藓(Plagiomnium maximoviczii(Lindb.)Kop.)、大凤尾藓(Fissidensnobilis Griff.)、大羽藓(Thuidium cymbifolium(Doz.et Molk.)B.S.G.)和大灰藓(Hypnum plumaeforme Wils.)嫩茎和老茎的石蜡切片和显微观察发现,同一藓类植株的嫩茎和老茎,茎结构稳定,不同种藓类植物茎横切面具有不同特征.植物体茎横切面形状、表层细胞的层数、细胞大小和细胞壁厚薄、皮层细胞大小和形状、中轴的有无以及比例等特征可以作为藓类植物的分科分类依据之一.     

Cause of Senescence of Nine Sorts of Flowers

The levels of endogenous phytohormones and respiratory rate in nine sorts of flowers such as Cymbidium faberi Rolfe, Nopalxochia ackermannii Kunth and others were investigated both at full bloom and senescence and meanwhile the effect of exogenous phytohormones on prolonging the blossoms and promoting ethylene production were tested. There is a high content of endogenous ethylene in all the long-lived flowere, about 3–16 folds higer than the short-lived ones. There is a high level of ABA at full blooming flowers of short-lived flowers, in which there is no or only some cytokinins in it, but the ratio of CTK (6BA+zeatin)/ABA is smaller(l.7). The endogenous ABA reached a much higher level at senescence in all nine sorts of flowers, so it is reasonable to consider that it is ABA which plays an important role of regulation in controlling flower's senescence. There is a much higher level of GA3 and zeatin in the long-lived flowers which is not demonstrated in the shortlived ones. The respiratory rate is one of the factors controtling the longevity of flowers, but it does not play a decided role. Application of 6BA and zeatin prolongs distinctly orchid’s longevity, however exogenous IAA through the promotive action on ethylene production, evidently extends the longevity of the flowers of the Nopalxochia ackermannii Kunth.     

龙胆科药用植物化学成分的研究现状

龙胆科植物在我国的分布范围很广,且多数为药用植物,其多数种属的药用植物,至今其化学成分尚未被系统研究。综述了目前龙胆科药用植物的化学成分的研究现状及一般提取方法,对近年来发现的环烯醚萜及裂环烯醚萜类化合物进行了总结,为本科药用植物的更深入研究提供了参考。     

The effects of glutamine of the maintenance of embryogenic cultures of Cryptomeria japonica

Summary Embryogenic tissues of sugi (Cryptomeria japonica) were induced on a modified Campbell and Durzan (CD) medium containing 1 μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 600 mg l⁻¹ glutamine, and subcultured in the medium of the same composition for over 1 yr. This resulted in a mixed culture of embryogenic and non-embryogenic cells. When embryogenic cells were isolated and cultured independently, their capacity to form embryogenic aggregates was lost. Thus, the non-embryogenic cells present within a mixed culture system were essential to the formation of embryogenic aggregates. When embryogenic tissues were isolated and cultured independently on a high glutamine-containing (2400 mg l⁻¹) medium, dry weights and endogenous levels of glutamine increased, and the tissue could generate a large number of embryogenic aggregates. Amino acid analysis of embryogenic and non-embryogenic cells from the maintenance culture indicated a higher level of glutamine was present in the latter. The high endogenous level of glutamine in the non-embryogenic portion of mixed cell masses may be the supplier of glutamine for maintaining the embryogenic property of the tissues.     

Specificity of action of piperidylcholine derivatives as substrates of cholinesterases of various origin

The review deals with study of enzymologic properties of a novel highly specific acetylcholinesterase substrate, N-(β-acetoxyethyl) piperidinium iodomethylate (“piperidylcholine”), and its 30 derivatives that were tested as effectors of cholinesterases of mammals and various species of Pacific squids. It was proven for the first time that responsible for specificity of action was structure of cyclic ammonium grouping of the alcohol part of molecule of the ester substrate. Analysis of specificity is performed based on enzymatic hydrolysis parameters—activity of catalytic center of cholinesterases and bimolecular constant of the reaction rate that are determined at optimal and low substrate concentrations. Among the specially synthesized group of thioester compounds there is revealed one more highly specific acetylcholinesterase substrate—N-(β-acetoxyethyl) piperidinium.     

真菌类遗传学分析的知识结构教学

本文以认知结构理论为指导,讨论了真菌类遗传分析与高等动植物遗传分析的内在联系,认为利用这种内在联系进行教学可收到好的效果并说明了作者的具体教学过程。 Abstract:In the paper, the relationship between genetic analysis of Fungi and genetic analysis of high animal and plant was discussed.A good results were obtained when we adopted this method in the teaching.     

医疗机构合理用药评价模型的构建研究

目的 针对医疗机构的合理用药水平进行评价研究。方法 根据医疗机构合理用药的具体要求,构建医疗机构合理用药评价指标体系,采用基于模糊群决策的方法和多指标评价分析法构建医疗机构合理用药评价模型。结果 构建了基于模糊群决策的医疗机构合理用药评价模型,并通过实例分析证明了评价模型的可行性。结论 建立的基于模糊群决策的医疗机构合理用药评价模型能够对医疗机构的合理用药水平进行科学评价,为提高医疗机构合理用药水平奠定基础。     

The mechanism of hatching of eggs of Haemonchus contortus

Fluid collected from hatching eggs of Haemonchus contortus contained a lipase which hydrolysed 2-naphthyl laurate (about 0·7 μmol naphthol freed /h/10⁶ eggs). The fluid also hydrolysed l-leucinamide (about 2·3 μmol leucine freed/h/10⁶ eggs). The fluid when added to normal or heated eggs caused ‘hatching’. ‘Hatching’ also occurred in exsheathing fluid from infective juveniles and in a preparation of pancreatic lipase containing leucine aminopeptidase. A purified mammalian leucine aminopeptidase in combination with several different lipases did not attack egg shells.The ‘spontaneous’ hatching of eggs of H. contortus was strongly inhibited by 1,10-phenanthroline, 10^?3M, and this inhibition was reversed by Zn²⁺. However, the inhibition of ‘hatching’ of eggs in externally applied hatching fluid, or the hydrolysis of leucinamide in hatching fluid was generally less marked.