Analysis of case histories on deep excavations in marine clay

Two case histories on deep excavation of marine clay are used to study the use of a decision-making tool based on a new deign method called the Mobilized Strength Design (MSD) method which allows the designer to use a simple method of predicting ground displacements during deep excavation. This application can approximately satisfy both safety and serviceability requirements by predicting stresses and displacements under working conditions by introducing the concept of "Mobilizable soil strength". The new method accommodates a number of features which are important to design of underground construction between retaining walls, including different deformation mechanism in different stages of excavation. The influence of wall depth, wall flexibility and stratified ground are the major focus of this paper. These developments should make it possible for a design engineer to take informed decisions on the influence of wall stiffness, or on the need for a jet-grouted base slab, for example, without having to conduct project-specific Finite Element Analysis.

Distinctive Paleo-Indian Migration Routes from Beringia Marked by Two Rare mtDNA Haplogroups

Background: It is widely accepted that the ancestors of Native Americans arrived in the New World via Beringia approximately 10 to 30 thousand years ago (kya). However, the arrival time(s), number of expansion events, and migration routes into the Western

鲢(Hypophthalmichthys molitrix)IL-10基因的克隆及表达分析

白细胞介素10(IL-10)是一种作用广泛的抗炎细胞因子,主要功能是限制和最终终止炎症反应．用RACE(rapid amplification of cDNA ends)-PCR方法扩增出鲢白细胞介素10(IL-10)的 cDNA,其全长为1248nt,包含5’非编码区156 nt,3’非编码区552nt,开放阅读框540nt．鲢IL- 10的开放阅读框编码179个氨基酸,其中包含构成两对二硫键的4个保守半胱氨酸．RT-PCR结果显示鲢IL-10 mRNA主要在脾脏、鳃、头肾和肌肉中表达．将鲢IL-10完

鳜IL-1β基因的克隆及原核表达

用RACE PCR获得了鳜白介素-1β(IL-1β)的全长cDNA.鳜IL-1β的cDNA全长为1298 nt(核苷酸),其5′非编码区包含UTR 93 nt;3′非编码区包含452 nt;其开放阅读框内包含753 nt,翻译成251个氨基酸.将鳜IL-1β克隆到原核表达载体pET-32a上,在大肠杆菌Rosetta- gami(DE3)内以包涵体形式得以高效表达.

草鱼、中华鳖脾细胞培养上清液IL-2物质的检测

用刀豆蛋白A(ConA)刺激诱导草鱼和中华鳖脾细胞 ,收集细胞培养上清液 ,用小鼠胸腺细胞增殖试验和对小鼠L92 9细胞系杀伤试验检测上清液中白细胞介素 2 (简称IL 2 )活性。结果表明 :草鱼、中华鳖脾细胞培养上清液中有IL 2样活性物质 ,这种物质使小鼠的胸腺细胞3H TdR掺入量明显增加 ,在相同效靶比的条件下对小鼠L92 9细胞系杀伤率也显著增强 ,这种IL 2活性均能被抗人rIL 2血清所抑制。中华鳖脾细胞培养上清液 (含IL 2 )对中华鳖胸腺细胞也有较明显的促增殖作用并能消除兔抗中华鳖胸

草鱼、中华鳖IL-2活性影响因素的研究

结果显示 :当年草鱼种和中华鳖脾细胞培养上清液中能检测出IL 2活性 ,而且 1龄以上草鱼、中华鳖的IL 2活性高于当年孵化的草鱼和中华鳖。草鱼、中华鳖脾细胞在 2 5℃培养温度条件下 ,其上清液中IL 2活性最高 ,35℃次之 ,1 5℃最低。因此 ,草鱼、中华鳖IL 2活性在一定范围内是随着年龄增加而增强和依赖温度的。通过小鼠胸腺细胞增殖和对小鼠L92 9细胞杀伤率的实验表明 37℃比 2 5℃检测温度下所测的IL 2活性高 ,而中华鳖胸腺细胞增殖实验却显示 2 5℃检测温度下IL 2活性高于 37℃

Molecular characterization and expression profiles in response to bacterial infection of Chinese soft-shelled turtle interleukin-8 (IL-8), the first reptilian chemokine gene

In this study, an IL-8 homologue has been cloned and identified from a reptile, Chinese soft-shelled turtle for the first time. The full-length cDNA of turtle IL-8 was 1188 bp and contained a 312 bp open reading frame (ORF) coding for a protein of 104 amino acids. The chemokine CXC domain, which contained Glu-Leu-Arg (ELR) motif and four cysteine residues, was well conserved in turtle IL-8. The 4924 bp genomic DNA of turtle IL-8 contained four exons and three introns. Phylogenetic analysis showed that the amino acid sequence of turtle IL-8 clustered together with birds. RT-PCR analysis showed that turtle IL-8 mRNA was constitutively expressed liver, spleen, kidney, heart, blood and intestine tissues of control turtles. Real-time quantitative PCR analysis further indicated that the turtle IL-8 mRNA expression was apparent in various tissues at 8 h and up-regulated significantly during 8 h-7 d after Aeromonas hydrophila infection. The present studies will help us to understand the evolution of IL-8 molecule and the inflammatory response mechanism in reptiles. (C) 2009 Elsevier Ltd. All rights reserved.

alpha-tocopherol is essential for acquired chill-light tolerance in the cyanobacterium Synechocystis sp strain PCC 6803

Unlike Escherichia coli, the cyanobacterium Synechocystis sp. strain PCC 6803 is insensitive to chill (5 degrees C) in the dark but rapidly losses viability when exposed to chill in the light (100 mu mol photons m(-2) s(-1)). Preconditioning at a low temperature (15 degrees C) greatly enhances the chill-light tolerance of Synechocystis sp. strain PCC 6803. This phenomenon is called acquired chill-light tolerance (ACLT). Preconditioned wild-type cells maintained a substantially higher level of alpha-tocopherol after exposure to chill-light stress. Mutants unable to synthesize alpha-tocopherol, such as slr1736, slr1737, slr0089, and slr0090 mutants, almost completely lost ACLT. When exposed to chill without light, these mutants showed no or a slight difference from the wild type. When complemented, the slr0089 mutant regained its ACLT. Copper-regulated expression of slr0090 from P-petE controlled the level of et-tocopherol and ACLT. We conclude that alpha-tocopherol is essential for ACLT of Synechocystis sp. strain PCC 6803. The role of a-tocopherol in ACLT may be based largely on a nonantioxidant activity that is not possessed by other tocopherols or pathway intermediates.

IL-2基因的突变及其与肠毒素融合基因的克隆和表达

由金黄色葡萄球菌分泌到胞外的单亚基蛋白中有肠毒素A和B。近年来两种毒素在肿瘤治疗研究方面取得了很大进展。白介素-2作为靶向分子，在抗肿瘤的药物中很有应用前景。本文分别对肠毒素A、肠毒素B和白介素-2进行了克隆和表达，并将白介素-2（125Ala）分别与肠毒素A227AI。肠毒素B进行了融合表达。从筛选到的天然金黄色葡萄球菌STSw的基因组中，通过PCR方法扩增出seb基因，同时突变了该基因两端几个稀有密码子。并将其克隆到7ZTS载体上进行表达，表达量占细胞总蛋白的33.5％。以seam基因为模板，通过重叠PCR将其227位天冬氨酸突变为丙氨酸，以降低其毒性。该突变基因重组到7ZTS载体中，并在JM109（DE3）中表达，表达量占细胞总蛋白的51.5％。通过重叠PCR法，对人的IL-2基因进行定点突变。共突变60个碱基，涉及51个氨基酸，其中第125位的半肤氨酸被突变为丙氨酸。该基因在大肠杆菌中表达量占总蛋白的30％。分别对上述三个工程菌的表达条件进行了探索。先制备出融合基因，再对融合基因进行表达，得到两种融合蛋白。它们是以6个甘氨酸和1个苏氨酸为链，将IL一2（125Ala）与肠毒素A227（Ala）、B分别连接起来，即IL-2（125Ala）-SEA227（Ala）和IL-2（125Ala）-SEB二者在大肠杆菌中表达量分别占总蛋白的10％和12％。以上实验结果为将几种蛋白开发成抗肿瘤靶向药物奠定了坚实基础。

IL-2与金黄色葡萄球菌肠毒素A和B融合基因的克隆及表达

目的 :IL - 2与金黄色葡萄球菌肠毒素A和B融合基因的克隆及表达。方法 :分别在金黄色葡萄球菌肠毒素A2 2 7Ala、B基因的两端克隆上两个酶切位点HindⅢ ,KpnⅠ。将IL - 2基因突变 ,设计一段linker使之分别与SEA2 2 7Ala和SEB相连并克隆到PET表达载体中 ,在大肠杆菌DH5α(DE3) -Pass中表达。结果 :表达的蛋白占总蛋白 15 %。结论 :IL - 2与金黄色葡萄球菌肠毒素A和B融合蛋白能在大肠杆菌中有效表达

定点突变提高IL-4免疫毒素对淋巴瘤的选择性和杀伤活性

采用软件分析选择与IL-4分子结合与活性相关的重要位点13T,121R,通过定点突变得到IL-4突变基因cpIL4(13D121E),将其与绿脓杆菌外毒素突变基因PE38KDEL融合,成功地构建了编码免疫毒素cpIL4(13D121E)-PE38KDEL的融合基因.该基因在原核表达系统中得到了高效表达,表达量占细胞全蛋白的30%以上.表达产物经亲和色谱和阴离子交换色谱纯化后,进行细胞毒性实验,证明其对表达型IL-4受体的淋巴瘤细胞Daudi具有良好的细胞毒作用,活性是同类型IL-4免疫毒素的2倍,而对表达型IL-4受体的内皮细胞活性较低.

The use of CE-electrochemiluminescence with ionic liquid for the determination of bioactive constituents in Chinese traditional medicine

CE/tris(2,2-bipyridyl) ruthenium(ll) (Ru(bpy)(3)(2+)) electrochemiluminescence (ECL), CEECL, with an ionic liquid (IL) detection system was established for the determination of bioactive constituents in Chinese traditional medicine opium poppy which contain large amounts of coexistent substances. A minimal sample pretreatment which involves a one-step extraction approach avoids both sample loss and environmental pollution. As the nearby hydroxyl groups in some alkaloid such as morphine may react with borate to form complexes and IL, as a high-conductivity additive in running buffer, could cause an enhanced field-amplified effect of electrokinetic injection. Running buffer containing 25 mM borax-8 mM 1-ethyl-3-methylimidazolium tetrafluoroborate (EMImBF(4)) IL (pH 9.18) was used which resulted in significant changes in separation selectivity and obvious enhancement in ECL intensities for those alkaloids with similar structures. Sensitive detection could be achieved when the distance between the Pt working electrode and the outlet of separation capillary was set at 150 mu m and the stainless steel cannula was fixed approximately 1 cm away from the outlet of the capillary. Quantitative analysis of four alkaloids was achieved at a detection voltage of 1.2 V and a separation voltage of 15 kV in less than 7 min.

Molecular cloning and expression of interleukin 1beta (IL-1 beta) from red seabream (Pagrus major)

The interleukin 1beta (IL-1beta) cDNA was cloned from the red seabream (Pagrus major) by homology cloning strategy. A cDNA fragment was amplified by PCR using two degenerated primers, which were designed according to the conserved regions of other known IL-1beta sequences, and elongated by 3' ends and 5' ends RACE PCR to get the full length coding sequence of red seabream IL-1beta (RS IL-1beta). The sequence contained 1252 nucleotides that included a 5' untranslated region (UTR) of 84 bp, a 3' UTR of 410 bp and an open reading frame (ORF) of 759 nucleotides which could be translated into a putative peptide of 253 amino acids with molecular weight of 28.6 kD and putative isoelectric point pI of 5.29. The deduced peptide contained two potential N-glycosylation sites and an identifiable IL1 family signature, but lacked the signal peptide and the clear ICE cut site, which were common in other nonmammalian IL-1beta genes. The RS IL-1beta had the highest homology with piscine IL-1beta according to phylogenetic tree analysis. The transcript expression was detected in blood, brain, gill, heart, head kidney, kidney, liver, muscle and spleen in the pathogen challenged and healthy red seabream by RTPCR. Results showed that the RS IL-1beta mRNA was constitutively expressed in most of the tissues both in stimulated and un-stimulated fish, and the expression could be enhanced by pathogen challenging.

Identification and characterization of a myeloid differentiation factor 88 (MyD88) cDNA from Zhikong scallop Chlamys farreri

Myeloid differentiation factor 88 (MyD88) is a universal and essential adapter for the TLR/IL-1R family. In this report, the first mollusk Myd88 ortholog (named as CfMyd88) was cloned from Zhikong scallop (Chlamys farreri). The full-length cDNA of CfMyd88 was of 1554 bp, including a 5 '-terminal untranslated region (UTR) of 427 bp, a polyA tail, and an open reading frame (ORF) of 1104 bp encoding a polypeptide of 367 amino acids containing the typical TLR and IL-1R-related (TIR) domain and death domain (DD). Homology analysis revealed that the predicted amino acid sequence of CfMyd88 was homologous to a variety of previously identified Myd88s with more than 30% identity. The temporal expressions of CfMyd88 mRNA in the mixed primary cultured haemocytes stimulated by lipopolysaccharide (LPS) and peptidoglycans (PGN) were measured by real-time RT-PCR system. The mRNA expression of CfMyd88 decreased after stimulation with both LPS and PGN, and the lowest level was about 1/3 times (at 6 h) and 1/10 times (at 9 h) to that in the control group, respectively. The expression then recovered and was upregulated to two-fold at 9 h after LPS stimulation or to the original level at 12 It after PGN stimulation. The results suggest that the MyD88-dependent signaling pathway exists in scallop and was involved in the defense system. (c) 2007 Elsevier Ltd. All rights reserved.