小鼠淋巴细胞抗原CTLA-4的胞外肽段表达及纯化

目的: 真核细胞表达小鼠淋巴细胞抗原CTLA-4胞外段肽,研究表达肽段与抗原呈递细胞B7分子结合后减轻小鼠淋巴细胞刺激后的增殖抑制,从而启动T淋巴细胞进一步增殖。方法:从小鼠脾脏淋巴细胞获得总RNA,通过逆转录PCR扩增出CTLA-4全长基因,克隆并测序。依据胞外段序列和真核表达载体pcDNA3.1序列,合成引物扩增胞外片段,两者经内切核酸酶处理、连接构建重组表达pcDNA3.1载体,重组质粒经测序验证后,采用lipofectamine 2000转染入小鼠肝癌细胞Hepa1-6,经G418筛选获得稳定表达细胞株。结果:获得小鼠CTLA-4胞外段真核表达载体和小鼠肝癌细胞Hepa1-6稳定表达转染细胞株,制备了CTLA-4胞外肽段,经His标签抗体和小鼠CTLA-4抗体Western blot检测表达蛋白带均呈阳性。结论:获得CTLA-4胞外段肽,为进一步研究该肽的作用打下基础。     

天童常绿阔叶树种栲树生殖个体大小及其生殖构件特征

对浙江天童木荷-栲树林内的常绿阔叶树种栲树(Castanopsis fargesii Franch.)的生殖个体大小、生殖构件的分布及其动态变化特征进行了研究。结果表明, 该地区栲树生殖个体的胸径在17~50 cm 间, 平均胸径为31. 2±8.0 cm, 平均年龄约36.3±6.6 年;林缘附近的生殖个体小于木荷-林内。相对稳定的群落和比较丰富的土壤养分条件有利于生殖枝数量和花序数量的增多。栲树生殖个体的数量在两年中变化较大, 部分栲树个体可以在连续年份中生殖。从枝系水平分析:在持续生殖的栲树个体上, 生殖枝数量有明显变化, 并非所有的生殖枝在两年中都可开花或结果, 保持连续生殖的枝系约占48.2%。栲树果序枝数量在连续年份有明显差异(p < 0.01), 而且果序枝上的幼蕾数、果实数量及结实率等都有明显差异(p < 0.05)。     

骨软骨复合支架的研究进展

软骨的修复是当前医学界十分棘手的难题,人们采取若干手段均收效甚微。由于软骨缺损时,其下的软骨下骨常出现硬化、退变,而新生软骨是无法与病变的软骨下骨进行整合的,所以在修复软骨的同时,必须重视软骨下骨的修复。近十几年来,人们开始发明和利用各种骨软骨复合支架,进行同时修复软骨与软骨下骨的动物实验研究。在正常骨软骨组织中,软骨与软骨下骨被钙化层所相连,此外钙化层也将软骨与软骨下骨分隔在不同的生存环境中。根据仿生学原理,人们又设计出一种带有隔离层的新型骨软骨复合支架,并取得了较为理想的实验结果。本文就国内外骨软骨复合支架的研完进展作一综述。     

Preliminary crystallographic data for protease omega

Protease omega from Carica papaya L. has been purified and crystallized. The crystals are trigonal, space group P3(1)12 (or P3(2)12), with a = 7.42 +/- 0.02 nm, c = 7.79 +/- 0.02 nm with one molecule in the asymmetric unit. The crystals diffract to 0.19-nm resolution using synchrotron radiation.     

Membrane Skeletal Proteins and Their Integral Membrane Protein Anchors are Targets for Tyrosine and Threonine Kinases in Euglena

ABSTRACT. Proteins of the membrane skeleton of Euglena gracilis were extensively phosphorylated in vivo and in vitro after incubation with [³²P]-orthophosphate or γ-[³²P] ATP. Endogenous protein threonine/serine activity phosphorylated the major membrane skeletal proteins (articulins) and the putative integral membrane protein (IP39) anchor for articulins. The latter was also the major target for endogenous protein tyrosine kinase activity. A cytoplasmic domain of IP39 was specifically phosphorylated, and removal of this domain with papain eliminated the radiolabeled phosphoamino acids and eliminated or radically shifted the PI of the multiple isoforms of IP39. In gel kinase assays IP39 autophosphorylated and a 25 kDa protein which does not autophosphorylate was identified as a threonine/serine (casein) kinase. Plasma membranes from the membrane skeletal protein complex contained threonine/serine (casein) kinase activity, and cross-linking experiments suggested that IP39 was the likely source for this membrane activity. pH optima, cation requirements and heparin sensitivity of the detergent solubilized membrane activity were determined. Together these results suggest that protein kinases may be important modulators of protein assembly and function of the membrane skeleton of these protistan cells.     

上海城市近自然森林的重建动态

近自然森林作为城市植被恢复的重要模式,已在我国多地开展了实践,并衍生了新的造林模式。为评估不同近自然森林建设模式的群落恢复进程,本研究以上海城市裸地上重建的近自然森林为对象,通过长期监测两处分别应用原"宫胁法"与新"异龄复层落叶—常绿混交林"种植模式的近自然森林重建过程,从物种组成、垂直结构、生活型组成和目标恢复物种4个方面解析恢复动态。结果表明:两种造林模式恢复的近自然森林,随恢复进程其物种组成逐渐趋同,在十多年内已形成了落叶—常绿垂直混交结构;"异龄复层落叶—常绿混交林"造林模式可更好地促进常绿阔叶树种的恢复,尤其是常绿建群种红楠与小叶青冈。本研究证实了近自然森林恢复技术可以缩短亚热森林群落向演替后期发展的时间,以及新造林模式的有效性,为近自然森林技术的应用与实施提供了科学依据。     

Protein engineering to change thermal stability for food enzymes.

In this review we have briefly indicated how the present state of knowledge allows proteins to be mutated to increase or decrease stability. We have discussed experiments on both model proteins and those of relevance to the food industry, and show how hydrophobic forces are a major driving force for folding as well as having a major role in thermostability. We have also indicated the large contribution that hydrogen bonding, electrostatic interactions and, in a less well predicted way, disulphide bridges make to thermostability.     

Structure of the Chlamydomonas agglutinin and related flagellar surface proteins in vitro and in situ

Using the quick-freeze, deep-etch technique, we compare the structure of the cane-shaped plus and minus sexual agglutinin molecules purified from gametes of Chlamydomonas reinhardi. We also describe the structure of three additional gamete-specific fibrillar molecules, called short canes, loops, and crescents, which are structurally related to the agglutinins. Four non-agglutinating mutant strains are found to produce the three latter fibrils but not canes, supporting our identification of the cane-shaped molecule as the agglutinin. The heads of the plus and minus canes are shown to differ in morphology. Moreover, two treatments that inactivate the plus agglutinin in vitro--thermolysin digestion and disulfide reduction/alkylation--bring about detectable structural changes only in the head domain of the cane, suggesting that the head may play an indispensible role in affecting gametic recognition/adhesion. We also present quick-freeze, deep-etch images of the flagellar surfaces of gametic, vegetative, and mutant cells of Chlamydomonas reinhardi. The gametic flagella are shown to carry the canes, short canes, loops, and crescents present in in vitro preparations. The cane and crescent proteins self-associate on the flagellar surface into stout fibers of uniform caliber, and they align along the longitudinal axis of the flagellum. The short canes and loops co-purify with flagella but, in the presence of mica, dissociate so that they lie to the sides of the flagella. The agglutinin canes of both mating types are oriented with their hooks at the membrane surface and their heads directed outward, where they are positioned to participate in the initial events of sexual agglutination.     

Empirically testing the effectiveness of thermal imaging as a tool for identification of large mammals in the African bushveldt

Monitoring animal populations often relies on direct visual observations. This is problematic at night when spotlighting can cause misidentification and inaccurate counting. Using infrared thermography (IRT) could potentially solve these difficulties, but reliability is uncertain. Here, we test the accuracy of 24 observers, differing in experience and skill levels, in identifying antelope species from IRT photographs taken in the African bush. Overall, 38% of identifications were correct to species level, and 50% were correct to genus/subfamily level. Identification accuracy depended on the confidence and skill of the observer (positive relationship), the number of animals present (positive relationship), and the distance at which it was taken (negative relationship). Species with characteristic features, horn morphology, or posture were identified with ~80% accuracy (e.g. wildebeest, kudu and impala) while others were considerably lower (e.g. blesbok and waterbuck). Experience significantly improved identification accuracy but the effect was not consistent between species and even experienced observers struggled to identify red hartebeest, reedbuck and eland. Counting inaccuracies were commonplace, particularly when group size was large. We conclude that thermal characteristics of species and experience of observers can pose challenges for African field ecologists, but IRT can be used to identify and count some species accurately, especially <100 m.