白喉毒素-血管内皮生长因子突变体融合毒素的研究

本研究旨在利用突变后的血管内皮生长因子（VEGF）与白喉毒素（DT）构建靶向毒素，使其特异性地杀伤肿瘤血管，达到阻断血管增生，切断肿瘤的血液供应的目的。首先通过基因工程技术从白喉杆菌中提取基因组DNA，扩增出其中的DTC区、T区基因。并运用不同的突变方法，构建了四个VEGF的突变体。即VEGF R82A，K84A，H86A突变，VEGF D63A，E64A，E67A突变，和截去了肝素结合区和NP-l（neuropilin-l）受体结合区的VEGF R82A，K84A，H86A突变和VEGF D63A，E64A，E67A突变。以这些突变体的编码基因与DT的T区、C区基因制成四个融合基因，并在大肠杆菌中表达、纯化、复性。以相似条件下制备的不加VEGF突变体的DTT区、C区蛋白为阴性对照，以VEGFR-l＋和VEGFR-2＋的Lovo细胞和VEGFR-l＋VEGFR-2-的SMMC-7721细胞做细胞实验。制成的四个融合毒素中有三个通过细胞学检验，具有靶向杀伤作用。特异性结合VEGFR-1受体的蛋白PmV8D、PsmVSD既可以抑制VEGFR-1+、VEGFR-2-的肝癌细胞SMMC-7721又可以抑制VEGFR-1+、VEGFR-2＋的结肠癌细胞Lovo。而不带VEGF突变体的DT391无抑制作用。特异性结合VEGFRZ受体的蛋白PmV6D可以抑制VEGFR-1+、VEGFR-2＋结肠癌细胞Lovo的生长，但不可以抑制VEGFR-1＋，VEGFR-2-的肝癌细胞SMMC-7721。受体结合力过弱的PsmV6D不对细胞的生长产生任何影响。

杀伤血管内皮生长因子受体1阳性细胞的靶向毒素

白喉毒素(diphtheria toxin DT) 是棒状白喉杆菌被茁噬菌体感染后分泌的一种外毒素. 它可以阻断真核细胞的蛋白质合成,杀死细胞. 血管内皮生长因子(VEGF) 的R82A,K84A,H86A突变体可以和肿瘤血管上高表达的VEGF受体1 (VEGFR-1) 特异性结合. 首先从白喉杆菌中提取基因组DNA,扩增出白喉毒素C区、T区基因. 并运用点突变技术,制成VEGF的R82A,K84A,H86A突变体. 利用这个可以和肿瘤血管上特异性受体相结合的VEGF的突变体,代替白喉毒素上的受体结合区,制成了针对VEGFR-1的靶向融合毒素——DT391-mVEGF. 以去除了受体结合区的DT391为阴性对照,细胞实验表明,融合毒素对VEGFR-1阳性的肿瘤细胞有特异性杀伤作用.

Mitochondrial cytochrome b gene sequences of Old World monkeys: With special reference on evolution of Asian colobines

The classification and phylogenetic relationships of the Old World monkeys are still controversial. For Asian colobines, from three to nine genera were recognized by different primatologists. In the present study, we have sequenced a 424 bp mitochondrial tRNA(Thr) gene and cytochrome b gene fragment from Macaca mulatta, Mandrillus sphinx, Mandrillus leucophaeus, Semnopithecus entellus, Trachypithecus vetulus, T. johnii, T. phayrei, T. francoisi, Pygathrix nemaeus, Rhinopithecus roxellanae, R. bieti, R. avunculus, Nasalis larvatus, and Colobus polykomos in order to gain independent information on the classification and phylogenetic relationships of those species. Phylogenetic trees were constructed with parsimony analysis by weighting transversions 5 or 10 fold greater than transitions. Our results support the following conclusions: (1) the Old World monkeys are divided into two subfamilies; (2) that among the colobines, Colobus, the African group, diverged first, and Nasalis and Rhinopithecus form a sister clade to Pygathrix; (3) that there are two clades within leaf monkeys, i.e. 1) S. entellus, T. johnii, and T. vetulus, and 2) T, phayrei and T. francoisi; (4) that Rhinopithecus avunculus, R. roxellanae, and R. bieti are closely related to each other, and they should be placed into the same subgenus; (5) that Rhinopithecus is a distinct genus; and (6) that the ancestors of Asian colobines migrated from Africa to Asia during the late Pliocene or early Pleistocene.

Stability and molecular modeling of triplex DNA inhibiting DNA binding protein binding to the core promoter of hepatitis B virus.

Two three-dimensional structure models of the 21nt oligodeoxyribonucleotides, CPI (G3TG-2TGT2G5TG2TGT) and CP3 (TGTG2TGST2GTG2TG3), were constructed by InsightII (MSI) software in IRIS Indigo2 (SGI) workstation using the crystal structure of TAT tripler formation as the template. The initial structures subsequently were minimized by molecular mechanics. The final structures were believed as the dominant conformation. The results showed that the energy of CP1 is lower than that of CP3, and the former is more stable than the latter. Moreover, the results further proved that the 21nt oligodeoxyribo-nucleotide CP1 stably combines with the core promoter (Cp) fragment of hepatitis B virus (HBV) to form a tripler DNA, and CP1 specifically inhibits a specific cellular factor (DNA binding protein) binding to Cp fragment. These results indicated that specific repression of gene transcription of HBV DNA might be possible by tripler-formation DNA.

Construction and characterization of a novel recombinant single-chain variable fragment antibody against White Spot Syndrome Virus from shrimp

An antibody phage display library against White Spot Syndrome Virus (WSSV) was constructed. After four rounds of panning against WSSV, 192 out of 480 clones displayed WSSV binding activity. One of the positive clones, designated A1, had relatively higher activity specifically binding to WSSV A1-soluble, single-chain fragment variable (scFv) antibody has an affinity constant (K-aff) of 2.02 +/- 0.42 x 10(9) M-1. Dot blot assays showed that A1-soluble scFv could detect WSSV directly from shrimp hemolymph after 24-h feeding infection by WSSV. A1 scFv has potential for the development of a cheap, simple and sensitive diagnostic kit for WSSV in the field. (C) 2003 Elsevier Science B.V. All rights reserved.

Analysis of expression of the binary toxin genes from Bacillus sphaericus in Anabaena and the potential in mosquito control

Anabaena strains expressing the binary toxin genes of Bacillus sphaericus produce high larvicidal activity with living cells. Western blot analysis showed that the 51-kDa and 42-kDa toxin proteins were stable in Anabaena. When a DNA fragment upstream of the 51-kDa protein gene was deleted, the toxicity was reduced by over a hundred-fold, whereas deletions at the coding regions showed that the cooperation of the two proteins expressed in Anabaena is essential for the larvicidal activity. Outdoor tests showed that the genetically altered Anabaena could keep containers with natural water from being inhabited by Culex larvae for over 2 months.

Kinetic energies of C-n(+) (n <= 58) fragment ions produced by nanosecond laser impact on C-60

Laser-induced fragmentation of C-60 has been studied using a time-of-flight mass spectrometric technique. The average kinetic energies of fragment ions C-n(+) (n <= 58) have been extracted from the measured full width at half maximum (FWHM) of ion beam profiles. The primary formation mechanism of small fragment ion C-n(+) (n < 30) is assumed to be a two-step fragmentation process: C60 sequential decay to unstable C-30(+) ion and the binary fission of C-30(+). Considering a second photo absorption process in the later part of laser pulse duration, good agreement is achieved between experiment and theoretical description of photoion formation. (C) 2009 Elsevier B. V. All rights reserved.

Isospin effect of projectile fragment yields

Isospin asymmetry is very important in the nuclear equation of state (EOS), isotope yield from the projectile fragments can give information of the reaction process. In this paper projectile fragment yields are measured in the collision Ar-36,Ar-40 + Ni-64 at incident energy 50 MeV/u with different isospin asymmetry project Ar-36,Ar-40, data analysis, particle identification and event selection are described. Isotope yields are compared in these two reactions, and axe also compared with the empirical parametrization of fragmentation cross-section calculated by EPAX.

A recent shellfish toxin investigation in China

A shellfish toxin investigation along the Chinese coast has recently been conducted using both HPLC and mouse assay methods. The results showed that DSP was widely distributed in different shellfish species in China. 26 out of 89 samples had DTX1 (dinophysistoxin-1) or OA (okadaic acid) but the DSP content in most shellfish samples did not reach the regulatory limit for human consumption adopted in many countries (20 mu g/100 g soft tissue). PSP was also found in 5 out of 96 samples along the coast. One sample, Chlamys nobilis from Hong Kong contained high levels of PSP (320 mu g STX equivalent/100 g soft tissue), compared to the regulatory limit (80 mu g STX equivalent/100 g soft tissue). After the recent outbreak of red tide in Hong Kong waters, three further shellfish samples were collected within 40 days to investigate the impacts of this event, It was shown that high levels of PSP continued to exist in Hong Kong waters. This report provides the first report of DSP and PSP distribution along the Chinese coast. (C) 1999 Elsevier Science Ltd, All rights reserved.

Effects of heat-treatment processes on the electroless Ni-B coating and its natural aging mechanism

A Ni-B coating was prepared with EN using potassium borohydride reducing agent. The as-plated micro-structure of the coating was confirmed from XRD to be a mixture of amorphous and supersaturated solid solution. Three kinds of phase transformation were observed from the DSC curve. Different from the previous works, the formation of Ni4B3 and Ni2B was found during some transformation processes. The key factors which influence the variation of micro-hardness and micro-structure in deposits are the formation, the size and amount of Ni3B, Ni4B3 and Ni2B. Aging of the deposits treated under some heat treatment conditions occurred at room temperature. Changes of the micro-hardness indicated aging phenomena evidently. the natural aging phenomena are concerned with various kinds of decomposition of borides, especially with Ni4B3 phase. The extent of natural aging depends on the formation and the quantity of Ni(4)B3 and Ni2B.

Direct numerical test of the B-G-K model equation by the DSMC method

In the present paper the rarefied gas how caused by the sudden change of the wall temperature and the Rayleigh problem are simulated by the DSMC method which has been validated by experiments both in global flour field and velocity distribution function level. The comparison of the simulated results with the accurate numerical solutions of the B-G-K model equation shows that near equilibrium the BG-K equation with corrected collision frequency can give accurate result but as farther away from equilibrium the B-G-K equation is not accurate. This is for the first time that the error caused by the B-G-K model equation has been revealed.

The application of B-P constitutive equations in finite element analysis of high velocity impact

We present in this paper the application of B-P constitutive equations in finite element analysis of high velocity impact. The impact process carries out in so quick time that the heat-conducting can be neglected and meanwhile, the functions of temperature in equations need to be replaced by functions of plastic work. The material constants in the revised equations can be determined by comparison of the one-dimensional calculations with the experiments of Hopkinson bar. It can be seen from the comparison of the calculation with the experiment of a tungsten alloy projectile impacting a three-layer plate that the B-P constitutive equations in that the functions of temperature were replaced by the functions of plastic work can be used to analysis of high velocity impact.

Quantifying Cell Binding Kinetics Mediated By Surface-Bound Blood Type B Antigen To Immobilized Antibodies

Cell adhesion is crucial to many biological processes, such as inflammatory responses, tumor metastasis and thrombosis formation. Recently a commercial surface plasmon resonance (SPR)-based BIAcore biosensor has been extended to determine cell binding mediated by surface-bound biomolecular interactions. How such cell binding is quantitatively governed by kinetic rates and regulating factors, however, has been poorly understood. Here we developed a novel assay to determine the binding kinetics of surface-bound biomolecular interactions using a commercial BIAcore 3000 biosensor. Human red blood cells (RBCs) presenting blood group B antigen and CM5 chip bearing immobilized anti-B monoclonal antibody (mAb) were used to obtain the time courses of response unit, or sensorgrams, when flowing RBCs over the chip surface. A cellular kinetic model was proposed to correlate the sensorgrams with kinetic rates. Impacts of regulating factors, such as cell concentration, flow duration and rate, antibody-presenting level, as well as pH value and osmotic pressure of suspending medium were tested systematically, which imparted the confidence that the approach can be applied to kinetic measurements of cell adhesion mediated by surface-bound biomolecular interactions. These results provided a new insight into quantifying cell binding using a commercial SPR-based BIAcore biosensor.

Recurrence plot analysis of DNA sequences

Recurrence plot technique of DNA sequences is established on metric representation and employed to analyze correlation structure of nucleotide strings. It is found that, in the transference of nucleotide strings, a human DNA fragment has a major correlation distance, but a yeast chromosome's correlation distance has a constant increasing. (C) 2004 Elsevier B.V All rights reserved.

基于B/S的海洋平台远程监控系统设计

本文提出基于浏览器/服务器（Browser/Server简写为B/S）海洋平台结构的远程监测控制系统,实现在客户端不间断对平台进行实时监视,获得现场采集数据,并以图形方式进行实时显示,从而为平台的安全运行和结构设计提供科学依据。