The intraspecific difference of the triose phosphate isomerase ( tim) gene from Giardia lamblia

目的　探讨蓝氏贾第鞭毛虫 (Giardialamblia)磷酸丙糖异构酶基因种内差异。方法　提取虫体总DNA ,对所有虫株磷酸丙糖异构酶 (tim)基因部分片段进行PCR扩增。测定序列后 ,用简约法和NJ法构建系统树进行系统发育分析。结果　共有 12 4个位点存在变异 (占所有测定序列中的 2 3% ) ,且大多数为发生在密码子的同义突变。两种构树方法所得二树的分枝结构相似 ,均将受试的 16株蓝氏贾第虫分为明显的两组。结论　宿主及地理因素对蓝氏贾第虫群体的遗传多样性影响不大。在DNA分子进化水平上 ,自然选择的影响十分显著。可将tim基因作为蓝氏贾第虫群体遗传结构一个十分有效的遗传标记。

蓝藻TIM上的一个特异抑制靶位点及其应用

本发明涉及蓝藻TIM上的一个特异抑制靶位点及其应用，属生物技术领域。该抑制靶位点位于蓝藻TIM Loop-6的铰链区，为蓝藻Synechocystis sp.TIM氨基酸编号为准的半胱氨酸位点-Cys176。该抑制靶位点可作为在设计水华抑制剂中的应用。

腐生型眼虫Astasia longa 两种TIM 同工酶cDNA的克隆和分析

　利用3′2 RACE 和5′2 RACE 技术, 从腐生型眼虫长变胞藻( Astasia longa) 克隆了磷酸丙糖异构酶 ( TIM) 的两个同工酶cDNA 全序列。分析表明: 它们分别编码定位于细胞质的胞质型TIM (cTIM) 和定位于质 体的质体型TIM (p TIM) ; 后者的N 端具有引导该酶定位到质体中去的典型“前导序列”。根据这些事实我们推 测腐生型眼虫A . longa 质体中可能存在功能性的TIM , 并进一步认为该质体可能不只是一般意义上的“叶绿体 退化的残迹”, 而仍是一种至少有TIM 参与其代谢活动的功能性细胞器[动物学报50 (3) : 414 - 419 , 2004 ] 。

蓝氏贾第虫分离株tim 基因序列分析比较研究

　为了探讨来源于不同地理位置贾第虫分离株之间的遗传学关系, 采用tim 基因扩增、限制性酶切和 DNA 序列分析方法对来自我国(C1、C2、、CH2、CH3)、柬埔寨(CAM )、澳大利亚(A 1、A 2) 和美国(BP、CDC) 共9 株 蓝氏贾第虫的基因型进行了分析比较。结果表明,A 1、A 2 和CAM 属第1 型(WB) ; CH2 和CH3 属第2 型(JH) ; C1、 C2、BP 和CDC 属第3 型(GS)。本实验结果提示, 虫株间遗传学关系并非由地理位置的远近所决定。来源于同一地 理位置的虫株其遗传学特性可有很大的差异。相反, 来源于地理位置相隔很远的虫株其遗传学特性却可极为相近。 贾第虫分离株基因型的确定可为本虫分子系统进化和分子流行病学研究提供重要资料。

Identification of triosephosphate isomerase (TIM) genes from Microcystis (Cyanobacteria : Chroococcales) and theoretical analyses of the potential of cyanobacterial TIM as a target for designing specific inhibitors

The genes encoding triosephosphate isomerase (TIM) in three species of Microcystis (M. aeruginosa, M. viridis and M. wesenbergii) were investigated. Reverse transcriptase-polymerase chain reaction indicated that they were transcribed in the cells. Analyses showed that their DNA and deduced amino acid sequences were highly conserved between all the three species, only a single nonsynonymous substitution was seen at position 31, from an Asp in M. aeruginosa and M. viridis to Glu in M. wesenbergii. Sequence alignment of these with 12 other known cyanobacterial TIM sequences showed that all the cyanobacterial TIMs had a very high level of amino acid identity (over 50% between each two). Comparison of the cyanobacterial TIMs with other reported TIMs (from diverse lineages of the three Domains) showed that they possessed common active-site residues and sequence motifs. All cyanobacterial TIMs have two common cysteine residues (Cys127 and Cys176), and the Cys176 is almost cyanobacteria-specific with only one exception in Streptomyces coelicolor. Both secondary structure alignment and comparative modelling of Synechocystis sp. TIM showed that Cys176 was located at the hinge region of the flexible loop-6 and might therefore be critical to the movement of TIM's loop-6, which is important to the function of the enzyme. Thus, the cyanobacterial TIM-specific Cys176 may be a potential site for the discovery of suitable drugs against cyanobacteria, and such drugs may have utility in controlling water blooms due to cyanobacteria.

几类处在关键进化地位藻类的TIM基因和糖酵解途径的研究

糖酵解作为细胞的重要的基本代谢途径广泛存在于各类生物中。但参与该途径的tim基因／酶和整个途径在细胞中的区室定位情况在一些处在关键进化地位的藻类中还存在许多未知或争议。本文首先对参与该途径的一个重要酶――磷酸丙糖异构酶（TIM）在两种不同营养方式的眼虫上进行了鉴定和序列结构分析，并结合了包括绿藻、红藻和动基体类等在内的其它生物的数据进行了分子系统分析；其次，对该途径在绿藻类的衣藻细胞中的区室定位情况进行了研究，并对该特殊的区室定位途径的进化进行了探讨。得到了如下结果和结论： 1）通过3’与5’-RACE实验，在光合型眼虫Euglena gracilis和Euglena intermedia和腐生型眼虫Astasia longa中各获得了一长一短两个TIM的全长cDNA序列，同时通过基因组DNA-PCR和序列搜索，还获得了两种绿藻和红藻的tim基因序列。前导序列分析显示眼虫的两个cDNA分别编码定位到胞质的cTIM和定位到叶绿体的cpTIM或质体的pTIM。腐生眼虫A. longa的pTIM与光合型眼虫E. gracilis的cpTIM高度相似，其前导序列也具有眼虫典型的核编码叶绿体蛋白质的典型特征；两类不同营养型眼虫的成熟TIM间的同源性达91.6％，并且具有其它TIM所共有的活性中心氨基酸和保守的序列motifs。这表明A. longa的pTIM确实是定位于质体、具有生物活性的cpTIM的同工酶，并提示A. longa的质体还具有与TIM相关的代谢功能（例如脂肪酸的合成），而不仅仅是一个叶绿体的“残迹”。 TIM的序列比对和分子系统分析结果显示：眼虫类和红藻类的TIM共有一个由两个氨基酸组成的插入；更重要的是，眼虫类TIM既没有与被认为与它共祖的动基体类的TIM聚在一起，又没有与被认为为其提供叶绿体（经二次内共生）的绿藻的TIM聚在一起，而是与红藻TIM聚为一枝。这提示眼虫与红藻间在进化历史上可能曾经有过基因交流。 2）采用生物信息学手段结合分子生物学实验，对衣藻基因组和转录组中参与糖酵解相关基因／酶进行了鉴定、定位预测和表达水平分析，结果表明：与其它极大多数的真核生物不同，衣藻细胞质中不具有完整的糖酵解途径，尽管该途径的后三步主要发生在胞质中，但前七步则是发生在叶绿体中的。 分子系统分析表明衣藻叶绿体中参与前6步和细胞质中参与最后2步的糖酵解酶都是胞质型起源，其中参与第4步的FBA更像是由很早时期的胞质型基因重复而来的；而第7步的PGK是由内共生形成叶绿体的蓝细菌的水平基因转移而来。这表明衣藻胞质中所缺少的糖酵解酶是一种次生性丢失的结果，而非原始的特征；其叶绿体中的糖酵解步骤应该是由于原有胞质型糖酵解酶的基因重复之后重新定位或者直接重新定位到叶绿体中，以及内共生产生叶绿体时由蓝细菌的水平基因转移所致。

“Deborah Numbers”, Coupling Multiple Space and Time Scales and Governing Damage Evolution to Failure

Two different spatial levels are involved concerning damage accumulation to eventual failure. nucleation and growth rates of microdamage nN* and V*. It is found that the trans-scale length ratio c*/L does not directly affect the process. Instead, two independent dimensionless numbers: the trans-scale one * * ( V*)including the * **5 * N c V including mesoscopic parameters only, play the key role in the process of damage accumulation to failure. The above implies that there are three time scales involved in the process: the macroscopic imposed time scale tim = /a and two meso-scopic time scales, nucleation and growth of damage, (* *4) N N t =1 n c and tV=c*/V*. Clearly, the dimensionless number De*=tV/tim refers to the ratio of microdamage growth time scale over the macroscopically imposed time scale. So, analogous to the definition of Deborah number as the ratio of relaxation time over external one in rheology. Let De be the imposed Deborah number while De represents the competition and coupling between the microdamage growth and the macroscopically imposed wave loading. In stress-wave induced tensile failure (spallation) De* < 1, this means that microdamage has enough time to grow during the macroscopic wave loading. Thus, the microdamage growth appears to be the predominate mechanism governing the failure. Moreover, the dimensionless number D* = tV/tN characterizes the ratio of two intrinsic mesoscopic time scales: growth over nucleation. Similarly let D be the “intrinsic Deborah number”. Both time scales are relevant to intrinsic relaxation rather than imposed one. Furthermore, the intrinsic Deborah number D* implies a certain characteristic damage. In particular, it is derived that D* is a proper indicator of macroscopic critical damage to damage localization, like D* ∼ (10–3~10–2) in spallation. More importantly, we found that this small intrinsic Deborah number D* indicates the energy partition of microdamage dissipation over bulk plastic work. This explains why spallation can not be formulated by macroscopic energy criterion and must be treated by multi-scale analysis.

Solid-state NMR study of bioactive binary borosilicate glasses

A series of binary borosilicate glasses prepared by the sol-gel method are shown to be bioactive. Tetraethyl orthosilicate (TEOS) and trimethylborate (TMB) in acidic medium are used to prepare xB(2)O(3)center dot(1-x)SiO2 glass systems for x = 0.045-0.167. The formation of a layer of apatite-like mineral on the glass surface becomes apparent after soaking in simulated body fluid for 48 h. We have measured the B-11-B-11 homonuclear second moments of the borosilicate glasses and inferred that no macroscopic phase separation occurred in our glasses. The B-11 chemical shift data also show that the formation of clustered boroxol rings is negligible in our glass system. Although the bioactivity of our borosilicate glasses is less than that of CaO-SiO2 sol-gel glasses, these simple binary systems could be taken as reference glass systems for the search of new bioactive borosilicate glasses. (C) 2008 Elsevier Ltd. All rights reserved.

马铃薯in vitro 块茎发育的研究及其遗传转化受体系统的建立

研究了BAP在SLM→TIM、MA→TIM和切段诱导系统中与in vitro块茎发育的关系。在SLM→TIM系统中BAP没有促进块茎发育的作用;在MA→TIM系统中BAP具有促进块茎发育的作用;切段培养在20 ℃、8小时光照条件下则其块茎发育对BAP有依赖。系统地研究了光周期、蔗糖浓度和外源细胞分裂素在促进切段块茎发育方面的作用及其交互影响。确定了由切段诱导块茎的最佳培养条件以及用于BAP吸收代谢研究的实验条件。讨论了切段诱导系统在理论研究上的价值和生产上应用的前景。研究了无菌苗、长切段及切段BAP吸收运转代谢特点及其与块茎发育关系。马铃薯植物系统对BAP的吸收运转是需能代谢过程。BAP在植物系统运转性差与其在组织的代谢特点有关。在块茎诱导早期有标物质在小块茎或匍匐茎末端积累，这可能促使细胞分裂从而诱发块茎发育;但在成熟块茎中放射性物质浓度很低。外源细胞分裂素在切段或匍匐茎局部积累不是块茎发育的充分条件。在有利于块茎发育的条件下代谢早期BAP活性代谢产物含量明显地高于对照，代谢一定阶段后二种处理的切段BAP代谢谱趋于接近，这一代谢特点与BAP促进块茎发育的生理效应有关。运用同位素示踪技术研究了切段蔗糖吸收特点，外源细胞分裂素促进蔗糖在切段系统积累从而诱导体细胞储藏组织生化分化。短日照（黑暗）、较高浓度蔗糖、合适浓度BAP都有促进离体切段块茎发育作用，这几个因子在提高切段诱导水平方面具有协同效应。诱导早期切段中有块茎专一性糖蛋白Patatin痕量的存在。当切段发育了较大块茎后（诱导约15天）切段中Patatin含量明显增加。诱导3天切段系统中即有Patatin mRNA高水平地转录。诱导7天阶段Patatin mRNA含量迅速下降。块茎发育很可能是通过几种激素（包括块茎诱导因子）协同地对植物系统同化物尤其是蔗糖源库关系的调节而实现的。 建立了向马铃薯植物引入外源基因的受体系统。叶园盘与农杆菌（pGV2260:: pGV941）共培养。转化植株在选择培养基上培养大约3周即可从愈伤组织或直接从叶片边缘产生。在含有高浓度卡那霉素的培养基上转化植株大多表型正常，切段能够发育块茎，叶片能够形成愈伤组织。转化再生植株均含有NPT－II活性而未转化Desiree无菌苗没有NPT－II活性。Southern分析表明NPT－II基因已整合入转化植物基因组中。这一实验系统的建立为向马铃薯植物引进具有重要经济价值的外源基因创造了条件。

基于磷酸丙糖异构酶基因序列的蓝氏贾第鞭毛虫分子系统发育的研究

[目的 ]探讨蓝氏贾第鞭毛虫种内系统发育及遗传多样性。 [方法 ]对不同来源虫株的磷酸丙糖异构酶(tim)基因进行 PCR扩增、序列测定后 ,用简约法和 NJ法构建分子系统树进行系统学分析。 [结果 ]在所测序列中共有 12 4个位点存在变异 (2 3% ) ,且大多数为发生在第三密码子的同义突变 ,两种构树方法所得两树的分枝结构相似 ,均将受试的 16株蓝氏贾第鞭毛虫分为明显的两组。 [结论 ]tim基因可作为研究蓝氏贾第鞭毛虫群体遗传结构一个有效的遗传标记

Timeless与生物钟基因

综述了timeless基因的发现、多态性和重要功能。timeless是最先被发现的两个生物钟基因之一。生物钟的昼夜节律由PER、TIM、CLOCK和CYCLE 4个生物钟齿轮组成的正负反馈回路进行调节。其中TIM可以受光因子调控,它还可以与PER形成异二聚体,通过正负调控方式调节果蝇的昼夜节律行为。

The stripe-backed weasel Mustela strigidorsa: taxonomy, ecology, distribution and status

1. The stripe-backed weasel Mustela strigidorsa is one of the rarest and least-known mustelids in the world. Its phylogenetic relationships with other Mustela species remain controversial, though several unique morphological features distinguish it from congeners. 2. It probably lives mainly in evergreen forests in hills and mountains, but has also been recorded from plains forest, dense scrub, secondary forest, grassland and farmland. Known sites range in altitude from 90 m to 2500 m. Data are insufficient to distinguish between habitat and altitudes which support populations, and those where only dispersing animals may occur. 3. It has been confirmed from many localities in north-east India, north and central Myanmar, south China, north Thailand, north and central Laos, and north and central Vietnam. Given the limited survey effort, the number of recent records shows that the species is not as rare as hitherto believed. Neither specific nor urgent conservation needs are apparent.

DNA sequence analysis of the triose phosphate isomerase gene from isolates of Giardia lamblia

Objective To confirm the genetic relation between Giardia lamblia (G. lamblia) isolates from different geographic regions of China and other countries. Methods Genomic DNA were extracted from the trophozoites or cysts of Giardia lamblia. The triose phosphate isomerase (tim) gene was amplified using polymerase chain reaction (PCR) technique. PCR products were digested with endonuclease and sequenced. The data of sequencing were analyzed with the DNAstar software and compared with that of the isolates acquired from GenBank. Results Of nine isolates of Giardia lamblia from China (C1, C2, CH2 and CH3), Cambodia (CAM), Australia (A1 and A2) and America (BP and CDC), respectively, 3 (A1, A2 and CAM) fit into Group 1 (WB), 2 (CH2 and CH3) into Group 2, and 4 (C1, C2, BP and CDC) into Group 3 (GS). The results confirmed the genetic relatedness of G. lamblia isolates from all over the world. Conclusion Genotyping isolates of G. Lamblia provides important information for establishing the phylogenetic relationship or for the epidemiological evaluation of the spreading of this organism.