大肠杆菌海藻糖合成酶基因对提高烟草抗逆性能的研究

编码大肠杆菌海藻糖合成酶的otsA基因由农杆菌介导引入野生型烟草植株并在花椰菜花叶病毒启动子序列 (CaMV35S)控制下获得表达。蒸发光散射高效液相层析法测定海藻糖实验表明 ,转基因烟草能够合成海藻糖 ,合成量达 1 4μg g叶片湿重 ;转基因烟草表现为耐盐性生长、干燥失重缓慢等抗逆表型。说明海藻糖合成酶otsA基因的引入 ,改变了烟草的糖代谢途径 ,同时也提高了植物的耐盐碱、耐干旱特性。     

昆虫海藻糖及其合成酶基因的特性与功能研究进展

海藻糖广泛存在于细菌、真菌、昆虫、无脊椎动物和植物等大量生物中。它不仅可以作为昆虫的能量来源,而且在抗逆等方面起着重要作用。海藻糖合成酶(Trehalose-6-phosphate synthase,TPS)是海藻糖合成过程中的一个关键酶。目前细菌、真菌和植物中都已经被发现和克隆,但其不存在于哺乳动物中。海藻糖是昆虫的"血糖",主要通过海藻糖合成酶和海藻糖-6-磷酸脂酶(Trehalose-6-phosphate phosphatase,TPP)在脂肪体中催化合成。TPS基因所编码的蛋白序列一般都包含两个保守的结构域:TPS和TPP,分别对应着酵母中的Ots A和Ots B基因。昆虫海藻糖合成酶的基因表达和酶活性的变化与昆虫的多项生理过程有着密切的关系,海藻糖合成酶有可能成为控制害虫的新靶标。     

海藻糖及其在生物工程方面的应用

自海藻糖发现以来对其化学性质、生理功能、作用机理、代谢途径等已进行了较为深入的研究,其分子生物学的研究也渐渐兴起。研究表明海藻糖能提高生物体对干旱、低温、高温、pH、盐渍等逆境条件下的抗性。离体试验表明海藻糖能保护生物膜、蛋白质的结构并能保持逆境下的酶活性,同时,外源海藻糖同样对生物体有保护作用。由于海藻糖具有这些独特的生物学功能,它已在许多方面得以应用,可作为食品工业的一种添加剂和甜味剂,使干燥食品在得水后保持原有的色、香、味;也可作为医药工业的非特异性生物制品和生化药品保护剂,使其在常温下保存,从而降低运输与储存费用;另外,在农业研究中可利用现代分子生物技术培育表达海藻糖的转基因作物,提高农作物的抗旱、抗冻等抗逆性能。     

海藻糖代谢途径相关基因及生物工程

海藻糖(Trehalose)是一种由两个葡萄糖分子通过α,α-1,l糖苷键连接的非还原性双糖。最早的记录是在19世纪初期作为黑麦的麦角菌的一种成分而被描述,后来发现海藻糖广泛存在于微生物、动物和植物体内,特别是在那些能抗脱水作用的生物中起着重要作用。这些特殊生物具有在脱水条件下存活多年的性质,包括所谓的“复苏植物”(Selaginella lepidophylla)、某些咸水虾、线虫及面包酵母等。当它们体内99％的水分被去掉之后,仍保持着能在获水后迅速复活的能力^[1]。研究表明,海藻糖对于生物抗逆具有重要的保护作用。海藻糖的应用研究因此得到了人们的广泛关注和重视,目前海藻糖已被用作酶、其它蛋白、生物制品甚至移植器官的保护剂。海藻糖作为生物体对抗环境胁迫的重要应激保护物质,在不同生物中存在多种合成和分解代谢途径,相关基因已相继被克隆和分析。海藻糖合成、分解及其调控是生物抗逆的重要机制,其相关基因的研究也是海藻糖生物工程的重要基础。     

海藻糖的生物合成与分解途经及其生物学功能

海藻糖是一类在干旱、低温、热击或脱水等逆境环境下具有独特抗逆保护作用的二糖,广泛分布于藻类、细菌、真菌和动植物体内。近年来,随着对海藻糖研究的深入,海藻糖已经被广泛应用到食品、医药、化妆品和分子生物学研究等领域。该文简述了海藻糖在生物体内的代谢途径、生物学功能和研究进展,并对灭蚊真菌Pythiumsp.GY1938菌株海藻糖代谢酶基因的研究前景加以展望。     

昆虫几丁质合成及其调控研究前沿

几丁质合成与降解是昆虫最重要的生理过程之一。本文根据国外和作者自己的研究,综述了昆虫几丁质合成及其调控研究进展。昆虫几丁质的生物合成通路始于海藻糖,终止于几丁质,其中共有8个酶参与。目前研究最多的为海藻糖酶和几丁质合成酶。昆虫存在2个海藻糖酶基因和2个几丁质合成酶基因。可溶性海藻糖酶基因对昆虫表皮的几丁质合成影响更大,而膜结合海藻糖酶基因则主要影响中肠的几丁质合成。几丁质合成酶A主要负责表皮和气管几丁质的合成,而几丁质合成酶B则负责中肠围食膜的几丁质合成。目前,昆虫几丁质合成的调控途径主要有两种:利用RNAi技术和几丁质合成抑制剂。     

垫状卷柏海藻糖-6-磷酸合成酶基因的克隆及功能分析

海藻糖-6-磷酸合成酶(Trehalose-6-phosphate synthse, TPS)是植物海藻糖合成途径的关键酶, 在旱生卷柏等复苏植物对逆境胁迫应答中起重要作用。文章以我国特有旱生植物垫状卷柏(Selaginella pulvinata)为材料, 采用同源扩增与RACE技术相结合的方法克隆了海藻糖-6-磷酸合成酶基因SpTPS1, cDNA全长3 223 bp, 包括一个2 790 bp的开放阅读框, 推导的氨基酸序列与模式物种的海藻糖-6-磷酸合成酶具有较高的序列相似性, 催化活性中心保守位点基本一致。酵母功能互补实验证明, 用SpTPS1基因开放阅读框转化的海藻糖合成酶基因突变(tps1△)酵母菌株, 可恢复在以葡萄糖作为唯一碳源培养基上的生长, 说明垫状卷柏海藻糖-6-磷酸合成酶基因SpTPS1的编码蛋白具有生物活性, 可应用于植物抗逆性的转基因改良。     

灰树花海藻糖合成酶基因的克隆及其在大肠杆菌中的表达

海藻糖主要作用是作为生物体的结构组分、以及保护生物膜和保护蛋白质。在灰树花中 ,海藻糖在干重中所占比例最高可达到 1 5 %～ 1 7% ,说明灰树花合成海藻糖的能力很强。将灰树花海藻糖合成酶基因克隆 ,并在大肠杆菌表达系统里表达。表达量为 1 90mg L。通过活性测定 ,证明在大肠杆菌中表达的海藻糖合成酶具有酶活性 ,结合基因工程和酶工程方法 ,为合成海藻糖的研究提供了新的方向     

胰岛素受体基因InR调控褐飞虱海藻糖代谢研究

在昆虫中已发现成熟的典型胰岛素信号通路,但是其调控海藻糖代谢途径的机制还未清晰。为探讨胰岛素受体基因在褐飞虱海藻糖代谢平衡及其发育的调控作用,本文采用RNAi技术抑制胰岛素受体(InR)基因的表达,测定处理后海藻糖、糖原和葡萄糖含量及海藻糖酶活变化,并检测InR、类胰岛素多肽(Ilp)、海藻糖代谢途径中关键基因的表达。研究结果表明dsRNA注射后能够显著抑制Ilp和InR基因的表达;InR1低表达后72 h能够显著抑制3种糖类物质的含量;InR表达抑制后72 h可溶性海藻糖酶活性上升,而膜结合型海藻糖酶活性下降;当InR表达受抑制后3个海藻糖酶和2个海藻糖合成酶基因的表达都显著下降。这些结果说明InR能够影响海藻糖等糖类物质的平衡。从而为将来通过调控昆虫血糖平衡来控制害虫提供理论依据。     

改造大肠杆菌合成海藻糖途径以高效合成海藻糖

海藻糖是相容性溶质的一种,因其具有多种生物学功能,在食品、化妆品、药品以及器官移植等方面均有很广泛应用。然而近几年生产海藻糖主要集中在使用酶催化的方法,虽然这种方法的转化效率高,但是却存在着副产物的问题,难以得到高纯度的海藻糖产品,严重制约了海藻糖的应用。本文通过基因工程技术在大肠杆菌Escherichia coli中构建了海藻糖高效合成新途径,通过全细胞催化合成海藻糖。利用PCR技术在哈氏噬纤维菌Cytophaga hutchinsonii中克隆获得海藻糖双功能合成酶基因(tpsp),采用E.coli pTac-HisA高效表达载体,实现海藻糖双功能合成酶基因(tpsp)高效表达,利用高效表达菌株进行全细胞催化,将葡萄糖高效转化为海藻糖。结果表明C.hutchinsonii海藻糖合成酶基因(tpsp)在E.coli中成功实现表达,该酶能够在胞内将葡萄糖高效转化为海藻糖,并将其转运到胞外,实现海藻糖的高效率合成,海藻糖的产量提高到1.2 g/L,相对转化率为21%。当将此高产菌株在发酵罐中进行转化时,海藻糖的产量达到13.3 g/L,葡萄糖的相对转化率达到48.6%。采用C.hutchinsonii海藻糖合成酶基因高效表达并且应用于海藻糖全细胞合成催化在国内外尚属首次报道,海藻糖的转化率及产率都已达到文献报道最高水平,本研究为开拓海藻糖生产新技术奠定了基础。     

籼稻颖花分化与退化对不同播期温光的响应

试验以三系杂交籼稻‘五优308’和‘天优华占’以及常规籼稻‘黄华占’为材料,在大田条件下,设置10个播期,研究田间不同温光条件对籼稻生育期天数、颖花分化和退化数的影响.结果表明: 温度对3个籼稻品种生育期的影响比日照长度大,平均温度升高1 ℃,播种-穗分化始期天数平均减少1.5 d,而穗分化历期天数与光照和温度的关系均不密切.不同播期间每穗颖花数和颖花分化数存在显著差异.穗分化期间平均温度、最高温度和最低温度升高,有效积温增加,昼夜温差扩大,光辐射增强,有利于穗分化期干物质积累和颖花分化,各品种趋势一致.穗分化期间有效积温增加50 ℃,颖花分化数增加10.5朵,昼夜温差扩大1 ℃,颖花分化增加14.3朵,总光辐射量增加50 MJ·m^-2,颖花分化数增加17.1朵.颖花退化率与温度呈现二次项相关,极端高温或极端低温的自然条件不利于颖花形成,但低温天气对颖花退化的影响大于高温.温度低于临界温度,颖花退化率大幅增加,穗分化期临界积温为550～600 ℃,日平均温度为24.0～26.0 ℃,日最高温度为32.0～34.0 ℃,日最低温度为21.0～23.0 ℃.适宜高温、昼夜温差大、光照辐射强的自然条件利于颖花分化,并减少颖花退化.     

Esophageal and tympanic temperature responses to core blood temperature changes during hyperthermia

Esophageal, rectal, tympanic, and central blood temperature, i.e., pulmonary artery and aortic arch, were recorded in three patients during iatrogenic whole-body hyperthermia for the treatment of advanced malignant metastatic cancer. Aortic temperature closely followed changes in pulmonary arterial temperature, with an average delay time of 27 s. Esophageal temperature reflected quantitatively and more quickly (avg lag time, 80 s) the temperature changes in the pulmonary artery than tympanic membrane temperature. Tympanic temperature was consistently lower than the blood temperature of the heart during steady state. Therefore it is suggested that esophageal temperature is a preferable index of central blood temperature. Additionally, measurement of esophageal temperature can be made more easily and safely than tympanic membrane temperature.     

全缘叶绿绒蒿的花内热量来源和温度调节功能

为探究全缘叶绿绒蒿( Meconopsis integrifolia )的花内热量来源和温度调节功能,该研究选择在全缘叶绿绒蒿的巴朗山居群,对其进行遮阴及去瓣处理,并采用红外热像仪监测全缘叶绿绒蒿的花内微环境温度日变化及花器官温度,用环境温度计监测环境温度。结果表明:(1)太阳照射显著提高全缘叶绿绒蒿花内微环境温度和花器官温度,全缘叶绿绒蒿的热量主要来源于太阳辐射。花内微环境昼夜温差显著低于环境昼夜温差,全缘叶绿绒蒿的花具有温度调节功能。(2)白天环境温度较高时,太阳照射显著提高全缘叶绿绒蒿花内微环境温度,花瓣会降低花内微环境温度; 夜间环境温度较低时,花瓣闭合会提高花内微环境温度; 花瓣闭合运动降低了花内微环境昼夜温差,产生了保温效果。(3)在太阳照射下,花器官温度差异显著,雌雄蕊温度显著高于花瓣温度,且花器官温度由雌蕊柱头中心点向外递减,全缘叶绿绒蒿能有效调控花器官各部位的温度。综上认为,全缘叶绿绒蒿的花内热量来源于太阳辐射,主要通过花瓣闭合运动降低花内微环境昼夜温差并能在太阳照射下调节各花器官的温度实现温度调节功能。     

The necessity for low temperature treatment on the eggs of Emma Cricket,Teleogryllus emma and the method of increasing the hatching rate after low temperature treatment

The necessity for low temperature treatment of Emma Cricket Eggs, Teleogryllus emma, to dormancy breaking and the method of increasing hatching rate after low temperature treatment were investigated. Emma Cricket Eggs were treated by constant temperature, room temperature, low temperature, step‐down low temperature. As a result, constant temperature treatment was the lowest on hatching rate. The hatching rates of the rest of other temperature treatments were not different statistically (one‐way ANOVA). In other word, low temperature treatment was not necessary to dormancy breaking. But low temperature treatment was necessary to control individual number of Emma Cricket, Teleogryllus emma, on the mass indoor‐rearing system. And the step‐up temperature treatment was necessary to increase hatching rate after low temperature treatment.     

A test of biochemical symmorphosis in a heterothermic tissue: bluefin tuna white muscle

Selective brain cooling (SBC) is defined as a brain temperature cooler than the temperature of arterial blood from the trunk. Surrogate measures of arterial blood temperature have been used in many published studies on SBC. The use of a surrogate for arterial blood temperature has the potential to confound proper identification of SBC. We have measured brain, carotid blood, and rectal temperatures in conscious sheep exposed to 40, 22, and 5 degrees C. Rectal temperature was consistently higher than arterial blood temperature. Brain temperature was consistently cooler than rectal temperature during all exposures. Brain temperature only fell below carotid blood temperature during the final few hours of 40 degrees C exposure and not at all during the 5 degrees C exposure. Consequently, using rectal temperature as a surrogate for arterial blood temperature does not provide a reliable indication of the status of the SBC effector. We also show that rapid suppression of SBC can result if the animals are disturbed.     

Effect of temperature on the escape of charged polymer chain from a repulsive nanopore

The effect of temperature on the escape of a charged polymer chain from a repulsive nanopore is studied by Langevin dynamics simulations. The conformation properties of the charged polymer chain are dependent on the temperature as it is in random coil state at high temperature and in compact globule state at low temperature. The scaling behaviour between the escape time and polymer length is independent of temperature, while the escape time decreases with increasing the temperature in an exponential way. Different temperature-dependent behaviours are observed for the escape time in three temperature regimes: low temperature where the polymer is in compact globule state, intermediate temperature around the coil-to-globule transition temperature, and high temperature where the polymer is in the random coil state. We further find that, with a decrease in the temperature, the total number of moving steps for the escape increases sharply in the low-temperature regime while the time duration for each moving step increases sharply in the intermediate temperature regime.     

Selecting temperature for protein crystallization screens using the temperature dependence of the second virial coefficient

Protein crystals usually grow at a preferable temperature which is however not known for a new protein. This paper reports a new approach for determination of favorable crystallization temperature, which can be adopted to facilitate the crystallization screening process. By taking advantage of the correlation between the temperature dependence of the second virial coefficient (B(22)) and the solubility of protein, we measured the temperature dependence of B(22) to predict the temperature dependence of the solubility. Using information about solubility versus temperature, a preferred crystallization temperature can be proposed. If B(22) is a positive function of the temperature, a lower crystallization temperature is recommended; if B(22) shows opposite behavior with respect to the temperature, a higher crystallization temperature is preferred. Otherwise, any temperature in the tested range can be used.     

Temperature-Sensitive Behavior of Paramecium caudatum

SYNOPSIS. The effect of temperature on the behavior of swimming cells of Paramecium caudatum has been investigated by photographic analyses of their tracks in uniform temperature, in temperature gradient, or in temperature changing with time. When the cells were placed in the temperature gradient, the frequency of discontinuous directional changes of cells swimming toward the optimal temperature, the temperature of the culture, was much lower than that of the cells swimming in the opposite direction. This difference in the frequency of directional changes explained the observed accumulation of the cells at - the optimal temperature. When the temperature was suddenly changed toward the optimum, a transient decrease of the frequency of directional changes was observed and when the temperature was changed in the reverse direction, a transient increase of the frequency was noted. This transient response to the temperature change was the origin of the dependence of the frequency of directional changes on the swimming direction in the temperature gradient. Finally, the relation between the magnitude of the transient response and the rate of the temperature change was derived.     

新疆农田作物覆膜地温极值的时空变化

土壤温度的上限和下限值极大地影响农田作物的生长发育．根据对新疆农田2年度不同观测时刻、不同覆膜条件及不同作物的最高和最低地温的分析,可知2年度全生育期地温极值均发生在0cm位置．地温极值依观测时刻的不同而变动．覆膜最低温度始终高于裸地,说明覆膜对提高低温段土壤温度有明显作用．最低温度的日变化可用二次函数表达;14：00和20：00时最高地温可分别用深度的椭圆曲线和线性关系表达．不同条件下的地温极值和气温具有线性关系,其中裸地最低温度与气温的相关性高于覆膜;玉米最低温度与气温的相关性高于棉花;最高温度与气温的相关性比最低温度与气温的相关性差．     

Growth of Rhizobium japonicum Strains at Temperatures Above 27 degrees C

A study was conducted to examine the growth responses of different Rhizobium japonicum strains to increasing temperatures, determine the degree of variability among strains in those responses, and identify temperature-related growth characteristics that could be used to select temperature-tolerant strains. Each of 42 strains was grown in liquid culture for 96 h at 19 incubation temperatures ranging from 27.4 to 54.1 degrees C in a temperature gradient apparatus. Growth was estimated by measuring the change in optical density over time. Strains differed in their responses to increasing temperatures. Three characteristic temperatures were determined for each strain: the temperature giving the maximum optical density at 96 h (optimum temperature), the maximum temperature allowing a continuous increase in optical density during the 96-h period (maximum permissive temperature), and the maximum temperature allowing growth of the cultures after they were transferred to a uniform incubation temperature of 28 degrees C (maximum survival temperature). The three characteristic temperatures varied among strains and had the following ranges: optimum temperature, from 27.4 to 35.2 degrees C; maximum permissive temperature, from 29.8 to 38.0 degrees C; and maximum survival temperature, from 33.7 to 48.7 degrees C. Significant positive correlations were found between maximum permissive temperature and optimum temperature and between maximum permissive temperature and maximum survival temperature. Eight strains which had the highest maximum permissive temperature, optimum temperature, and maximum survival temperature were considered tolerant of high temperatures and were able to grow at temperatures higher than those previously reported for the most tolerant R. japonicum strains. The strains were of diverse geographical origin, but the response to high temperatures was not related to their origin. Evaluation of the temperature responses in pure culture may be useful in the search for R. japonicum strains better suited to environments in which high soil temperature is a limiting factor.