滇虎榛中的化学成分

滇虎榛中的化学成分叶海亚陈昌祥郝小江（中国科学院昆明植物研究所植物化学开放研究实验室,昆明６５０２０４）ＴＨＥＣＨＥＭＩＣＡＬＣＯＮＳＴＩＴＵＥＮＴＳＦＲＯＭＯＳＴＲＹＯＰＳＩＳＮＯＢＩＬＩＳＹＥＨａｉ－Ｙａ,ＣＨＥＮＣｈａｎｇ－Ｘｉａｎｇ,ＨＡ...     

组蛋白脱乙酰化酶—1保守氨基酸中组氨酸定点突变的建立

为了研究蛋白脱乙酰化酶－１（ＨＤＡＣ１）保守氨基酸中组氨酸的定点突变对其功能的影响,需要建立ＨＤＡＣ１保守组氨酸定点突变的突变子。在克隆野生型ＨＤＡＣ１ｃＤＮＡ的基础上,利用Ａｌ－ｔｅｒｅｄ ＳｉｔｅⅡ体外突变系统对ＨＤＡＣ１保守氨基酸中的３个组氨酸位点进行突变,并用全自动测序鉴定。结果分别获得了ＨＤＡＣ１的Ｈ１４０Ｆ、Ｈ１７８Ｆ、Ｈ１７９Ｆ的定点突变子,为进一步研究ＨＤＡＣ１保守氨基酸定点突变对     

Mg~（2＋）促进线粒体H~＋－ATP酶的重建机理──H~＋－ATP酶复合体亚基水平的研究

用专一性标记蛋白质巯基（－ＳＨ）的荧光探剂ａｃｒｙｌｏｄａｎ测定含Ｍｇ２＋的Ｆ０－ＡＴＰ酶或Ｆ０－ＯＳＣＰ－Ｆ１－ＡＴＰ酶的脂酶体的构象与无Ｍｇ２＋者明显不同,前者的蛋白质的－ＳＨ基团处于疏水性更强的微环境中;在有Ｍｇ２＋和ＯＳＣＰ同时存在下重建的Ｆ０－Ｆ１－ＡＴＰ酶脂酶体较无ＯＳＣＰ者表现更高的水解活力或膜电位,表明ＯＳＣＰ增强Ｍｇ２＋的促进作用,这进一步提示Ｍｇ２＋通过改变膜脂的物理状态促进线粒体Ｈ＋－ＡＴＰ酶重建的间接作用。这些实验结果,从线粒体Ｈ＋－ＡＴＰ酶复合体的亚基水平的相关性上,对于我们提出的Ｍｇ２＋通过改变膜脂的物理状态使之具有合适的流动性,诱导嵌入脂双层的Ｈ＋－ＡＴＰ酶复合体的Ｆ０的构象发生变化并传递至复合体的催化中心Ｆ１,从而使重建Ｆ１－Ｆ０－ＡＴＰ酶具有较适合的蛋白构象,表现较高的重建酶活性的假设提供了直接的实验证据,精确地阐明了Ｍｇ２＋促进线粒体Ｆ０－Ｆ１－ＡＴＰ酶重建作用的分子机理。     

抗旱基因HDCS1的植物表达载体构建

在克隆了二棱大麦第３组ＬＥＡｃＤＮＡ,抗旱基因ＨＤＣＳ１的基因上,将其连接于ｐＢ１１２１的ＣａＭＶ３５Ｓ启动子和ＮＯＳ终止子之间,,构建了ＨＤＣＳ１的植物表达载体ｐＢＨＣ,并进行了ＰＣＲ和酶切鉴定,为进行植物抗旱基因工程研究创造了条件。     

猪生殖—呼吸道综合征病毒（PRRSV）CH—1a株糖蛋白基因的遗传变异分析

新近发现的猪生殖－呼吸道综合征病毒（ＰＲＲＳＶ）是单股ＲＮＡ病毒,属于不久前成立的动脉炎病毒科,为了比较国内分离的ＣＨ－１ａ株与国外毒株的分子遗传学关系,扩增并克隆了ＰＲＲＳＶ ＣＨ－１ａ株糖蛋白基因ＯＲＦ２￣５,测定了其核苷酸序列。用序列分析软件分析了其编码产物的分子量、等电点、疏水性、抗原性和有关位点,并与国外毒株进行了序列比较和系统发育进化分析。结果表明：ＣＨ－１ａ株与ＶＲ－２３３２株尽管密     

HCV非结构蛋白(NS2/NS3)基因表达载体的构建及其一级结构分析

应用逆转录－聚合酶链反应（ＲＴ－ＰＣＲ）技术,从ＨＣＶ感染者血清中扩增编码ＨＣＶ病毒蛋白酶的ＮＳ２－ＮＳ３ｃＤＮＡ片段,在其５′和３′端分别引入ＥｃｏＲⅠ和ＸｂａⅠ限制性内切酶位点,定向克隆至真核表达载体ｐｃＤＮＡ３,构建重组载体ｐｃＤＮＡ－ＮＳ２３,重组表达载体经限制性内切酶消化鉴定．用ＳＰ６和Ｔ７通用引物对目的基因片断进行序列分析．序列同源性分析结果表明,与ＨＣＶ－Ｊ、ＨＣ－Ｃ２有高度的同源性,与ＨＣＶ－１、ＨＣＶ－Ｊ６、ＨＣＶ－Ｊ８同源性差,提示所克隆的基因属ＨＣＶⅡ型．该区内重要的功能位点如Ｚｎ２＋依赖性金属蛋白酶催化中心、丝氨酸蛋白酶催化中心等均高度保守     

单纯疱疹病毒I型扩增子系列载体的构建

我们先前已报道了构建成一种能在ＨＳＶ ｔｓＫ株辅助下进行复制和包装,并表达β－半乳糖苷酶基因ｌａｃＺ的ＨＳＶ－１扩增子质粒ｐＨＳＬ,以及它的应用。该质料中依次含有ＨＳＶ－１复制起点ｏｒｉＳ序列及ＩＥ６８启动子、ｌａｃＺ基因、ＳＶ４０ｐｏｌｙＡ、ＨＳＶ－１包装信号‘ａ’序列和大肠杆菌质粒骨架。然而,该质粒中的报告基因ｌａｃＺ无法用简单的酶切方法卸载下来,继而装入目的基因。本研究在此基础上,新构建了一     

高海拔地区植物的光合特性

高海拔地区植物的光合特性卢存福,贲桂英（中国科学院植物研究所,北京１０００４４）（中国科学院西北高原生物研究所,西宁８１０００１）ＰＨＯＴＯＳＹＮＴＨＥＴＩＣＣＨＡＲＡＣＴＥＲＩＳＴＩＣＳＯＦＰＬＡＮＴＳＡＴＨＩＧＨＡＬＴＩＴＵＤＥＳＬｕＣｕｎ－ｆｕ...     

骤冷与饥饿对小鼠肝脏影响的实验研究

为探讨饥饿及饥饿与骤冷对动物肝脏的影响,本实验用健康昆明种小鼠２５只,随机分成正常组５只,饥饿组１０只,饥饿后再予冷刺激组１０只（下称骤冷组）。采用组织化学及酶组织化学方法观察糖原（ＰＡＳ反应）、ＳＤＨ（琥珀酸脱氢酶）,ＬＤＨ（乳酸脱氢酶）,ＣｈＥ（胆碱酯酶）、Ｍｇ２＋－ＡＴＰａｓｅ（镁激活三磷酸腺苷酶）,ＡＣＰ（酸性磷酸酶）。结果提示：饥饿时肝脏ＰＡＳ反应,ＳＤＨ,Ｍｇ２＋－ＡＴＰａｓｅ、ＣｈＥ活性显著下降,而ＡＣＰ活性明显增强;饥饿后骤冷时ＰＡＳ（反应）、ＳＤＨ、ＣｈＥ更显著下降,而ＡＣＰ及Ｍｇ２＋－ＡＴＰａｓｅ活性反而增强。     

PCR与ELISA检测丙型肝炎病毒的比较

ＰＣＲ与ＥＬＩＳＡ检测丙型肝炎病毒的比较李京培王明丽史百芬陆应玉（安徽医科大学２３００３２）丙型肝炎病毒（ＨＣＶ）感染后,患者血清中可以检测其特异的核酸序列ＨＣＶ－ＲＮＡ和抗－ＨＣＶ等。目前国内临床大多数ＨＣＶ的实验室诊断主要依靠抗ＨＣＶ－ＩｇＧ检查,但急性期抗体检出率仅能达到６０％（３７．６～８２．８％）,部分病例可呈阴性反应［１］。说明用ＥＬＩＳＡ检测ＨＣＶ有相当部分漏检,不能发现早期患...     

中国水仙查尔酮合酶cDNA的克隆及序列分析(简报)

中国水仙系石蒜科水仙属多年生草本植物。其花枝多,花香浓郁,素有“凌波仙子”的美称。但水仙花色单一,影响其观赏价值。花色形成与植物体内的一类次级代谢产物类黄酮有关。查尔酮合酶(Chalcone synthase,CHS)是类黄酮合成途径中的一个关键酶,在植物体内它催化丙二酰基辅酶A的三个乙酸基和对羟苯丙烯酰辅酶A的一个乙酸基的缩合,产生柚配基查尔酮(naringenin)。此中心中     

中国水仙查尔酮合酶cDNA的克隆及序列分析（简报）

Chalcone synthase (CHS) is a key enzyme in the biosynthesis of all classes of flavonoids. The production of flower pigment is specifically regulated by the activity of CHS. We cloned the cDNA sequence of CHS-A gene from Narcissus by PCR and analyzed the coding sequence of gene. The result demonstrated that the sequence of the coding region was 1167bp, encoding a protein of 389 amino acid which was more than 80% homology with CHS of the other 8 plants, such as Nicotine abacus and Solana tuberosum.     

The chalcone synthase multigene family of Petunia hybrida (V30): differential,light-regulated expression during flower development and UV light induction

We have analysed the expression of the 8–10 members of the gene family encoding the flavonoid biosynthetic enzyme chalcone synthase (CHS) from Petunia hybrida. During normal plant development only two members of the gene family (CHS-A and CHS-J) are expressed. Their expression is restricted to floral tissues mainly. About 90% of the total CHS mRNA pool is transcribed from CHS-A, wheares CHS-J delivers about 10% in flower corolla, tube and anthers. Expression of CHS-A and CHS-J during flower development is coordinated and (red) light-dependent. In young seedlings and cell suspension cultures expression of CHS-A and CHS-J can be induced with UV light. In addition to CHS-A and CHS-J, expression of another two CHS genes (CHS-B and CHS-G) is induced in young seedlings by UV light, albeit at a low level. In contrast to CHS genes from Leguminoseae, Petunia CHS genes are not inducible by phytopathogen-derived elicitors. Expression of CHS-A and CHS-J is reduced to a similar extent in a regulatory CHS mutant, Petunia hybrida Red Star, suggesting that both genes are regulated by the same trans-acting factors. Comparison of the promoter sequences of CHS-A and CHS-J reveals some striking homologies, which might represent cis-acting regulatory sequences.     

Inhibition of flower pigmentation by antisense CHS genes: promoter and minimal sequence requirements for the antisense effect

Introduction of a constitutive antisense full-length chalcone synthase (CHS) cDNA gene in petunia can result in an inhibition of flower pigmentation. We have evaluated some of the factors which may be important for the effectiveness of an antisense CHS gene.Antisense CHS genes encoding half-length or quarter-length RNA complementary to the 3 half of CHS mRNA are able to affect flower pigmentation, while a gene encoding RNA complementary to the 5 half of CHS mRNA did not show phenotypic effects in transgenic petunia plants. We demonstrate that the RNA encoded by the latter gene has a much lower average steady-state level in leaf tissue than the RNAs encoded by the other antisense gene constructs. We have compared the CaMV 35S and endogenous CHS promoter strengths and intrinsic stabilities of sense and antisense CHS RNAs. From the data we conclude that the constitutive antisense CHS genes are not likely to provide an excess of antisense RNA compared to the CHS mRNA derived from the endogenous genes.Effective inhibition of flower pigmentation is also observed when the antisense CHS gene is under control of the homologous CHS promoter. The results indicate that the mechanism of antisense inhibition cannot solely operate via RNA duplex formation between sense and antisense RNA.     

Antisense chalcone synthase genes in petunia: Visualization of variable transgene expression

Summary The constitutive expression of an antisense chalcone synthase (CHS) gene in transgenic petunia plants results with high frequency in a reduced flower pigmentation due to a reduction in the CHS mRNA steady-state level in floral tissue. Here we show that this reduction is specific for CHS mRNA; chalcone flavanone isomerase (CHI) and dihydroflavonol reductase (DFR) mRNA steady-state levels are unaffected. However, in white floral tissue a severe reduction in CHI specific activity is found, accompanied by an altered signal for CHI protein on western blots. We find no correlation between the phenotypic effect of the antisense CHS gene and its chromosomal position. For some of the antisense CHS transformants the flower phenotype is highly variable. We demonstrate that pigmentation in these plants can be influenced by gibberellic acid and light, suggesting that the variable flower phenotype is caused by changes in physiological conditions during flower development. The results not only indicate that flower pigmentation in these plants reveals the variable expression of the antisense transgene, but also show that genomic sequences flanking the transgene may render its expression extremely susceptible to physiological conditions.     

Chalcone synthase gene lineage diversification confirms allopolyploid evolutionary relationships of European rostrate violets

Phylogenetic relationships among and within the subsections of the genus Viola are still far from resolved. We present the first organismal phylogeny of predominantly western European species of subsection Rostratae based on the plastid trnS-trnG intron and intergenic spacer and the nuclear low-copy gene chalcone synthase (CHS) sequences. CHS is a key enzyme in the synthesis of flavonoids, which are important for flower pigmentation. Genes encoding for CHS are members of a multigene family. In Viola, 3 different CHS copies are present. CHS gene lineages obtained confirmed earlier hypotheses about reticulate relationships between species of Viola subsection Rostratae based on karyotype data. Comparison of the CHS gene lineage tree and the plastid species phylogeny of Viola reconstructed in this study indicates that the different CHS copies present in Viola are the products of both recent and more ancient duplications.