Abstract:Caveolae are 50～100 nm membrane micro-invaginations associated with the plasma membrane of a variety of cells. They were first identified in transmission electron micrographs 40 years ago. Functionally, caveolae are thought to participate in transcellular transport of both small and large molecules across capillary endothelial cells. In addition, caveolae have been postulated to function in potocytosis. More recently, there is growing evidence that caveolae also participate in transmembrane signal transduction.

Abstract:On the basis of the transition-state theory, catalytic antibodies could be induced in body by using a transition-state analogue as hapten which was selected according to the specific reaction mechanism. A fuller description is given on the principle of hapten design for generation of catalytic antibodies, the relationship between catalytic antibodies and non-catalytic antibodies and the comparison of catalytic antibodies with enzymes. The application prospects of catalytic antibody in medicine science and the limitations in the application are also discussed.

Abstract:Recently, phage display techniques have been applied to various biological aspects. Phage can be exploited to display peptides, protein domains and proteins on its surface. Especially peptide phage display is now well established as a tool for the search for peptides binding to antibodies, enzymes, receptors, lectins, nucleic acids or other target molecules and in studying the substrate specificity of enzymes. It has great potential for application in the areas of drug discovery, vaccine development and other pharmaceutical development.

Abstract:Rapidly isolating transcriptional sequences from large fragment genomic DNA is a key step in the positional cloning of disease genes.Exon trapping is one of the most successful methods. The basic principle, methodology improvements and practical achievements of the exon trapping approach are reviewed in detail.

Abstract:The membrane proteins of synaptic vesicle and their role in the neurotransmitter release have been extensively studied. Recent progress has occurred in this field. Synapsin Ⅰ, synaptophysin, synaptoporin, synaptobrevin, synaptotagmin etc. are important proteins in membrane of synaptic vesicle. They play local and autonomous regulation in the docking, membrane fusion and exocytosis of synaptic vesicle, especilly in the mechanism of exocytotic fusion. A brief review of these aspects has been made.

Abstract:In addition to its well-known effects on metabolism, insulin stimulate the growth and proliferation of a variety of cells in culture, and evidence suggests that insulin may also be an important regulator of growth in vivo. The growth-promoting effects of insulin appear to be mediated by its own receptors, but in some cells, insulin appears to stimulate growth at high concentrations by activating insulin-like growth factor Ⅰ(IGF-Ⅰ) receptors. The post-receptor signal transduction consists of a series of receptor-associated tyrosine protein kinase activated intracellular protein phosphorylation and dephosphorylation reactions, such as the phosphorylation of insulin receptor substrate-1(IRS-1), Shc, Ras and phosphatidylinositol 3-kinase (PI3-K). It is supposed that some regions or sites on the insulin molecule may contribute more to the growth-promoting effects, and some preliminary evidences have come from the studies on several high mitogentic potent insulin analogues.

Abstract:Inter α trypsin inhibitor (ITI) is a kind of protein widely existed in human and animal blood. ITI was composed of three peptides convalently linked together by a chondroitin sulfate chain. Two of the three peptides were linked to the chondroitin sulfate chain each by an ester bond between the carboxyl group of the final amino acid of the peptide and an internal C-6 hydroxyl group of the sugar chain. In extracellular matrix, this kind of ester bond can be transferred from chondroitin sulfate chain to hyaluronan. Thus the extracellular matrix and cell functions were regulated. The other peptide chain of ITI has kunitz type protease inhibitor domains. As a medicine, it can be separated from urine.

Abstract:Ligation-mediated PCR is a strategy of single-sided PCR based on ligation. First, a common linker is linked to one end of DNA fragments, then DNA sequence is amplified between the common linker and a sequence-specific primer. The application of vent DNA polymerase and strategies of extension products capture and linker tag selection greatly improves the sensitivity of ligation-mediated PCR. The invention of this technique has facilitated the execution of in vivo footprinting study.

Abstract:Human prostate-specific antigen (PSA) gene has been shown to be regulated by androgen, and its androgen response element (ARE) has been located at about －170. To determine whether the androgen inducion of the gene was affected by upstream sequence of ARE, different natural and mutated DNA fragments of PSA promoter were linked to the CAT reporter gene respectively, and different pBLCAT3-PSA plasmids were constructed and used for transfection in human prostatic cancer cell line PC-3. The results indicated that a 15 bp segment of RF15 (－340～－326) could cooperate with ARE in maximal androgen induction significantly. The bandshift assay showed that some nucleic regulatory protein from human prostatic cancer cell line LNcap and PC-3 could bind to RF15 DNA fragment. And the protein ability to bind to RF15 was influenced by Zn²⁺. The results suggested that the RF15 may be a new regulatory sequence in the PSA promoter region. The regulatory protein, which binds to the RF15 sequence, may enhance the androgen induction via interaction with androgen receptor.

Abstract:The experiments were designed to test follistatin mRNA expression and regulation by EGF and GM-CSF in human choriocarcinoma cell line Jar using reverse-transcription polymerase chain reaction(RT-PCR) technique. The results showed as follows: 1. EGF induced a dose-dependent accumulation of follistatin mRNA level, with maximal increase at a dose of 1.0 nmol/L 2. Although GM-CSF did not affect the follistatin mRNA expression alone, it could reduce the follistatin mRNA level enhanced by EGF in a dose-dependent manner, the maximal effect dose was 10 nmol/L, the rate of inhibition could reach 62.3%. Those results indicated that follistatin gene expression was regulated by both hormones and growth factors in endocrine and paracrine/autocrine manner.

Abstract:After the fusion between the C₆₀-phosphatidylcholine liposome (C₆₀-concentration as 20 mg/L) and HeLa cells and illumination with the Tungsten-Halogen lamp (4 000 lx, 30 min), the most of cells were killed by the evaluation of MTT assay (3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide). Biochemical test showed that photoexcited C₆₀ led to the decrease of the sulfhydryl content of the membrane protein and the increase of the malonidaldehyde (MDA) with the peroxidation of the membrane lipid. SDS-PAGE showed the conjugation of the membrane protein. Fluorescence polarization (FP) indicated that the fluidity of the cell membrane increase.

Abstract:Ganglioside GM3 and bovine brain Gangliosides(BBG) were added exogenously to human glioblastma multiform cell line BT325 to observe their effects on BT325. The results showed that GM3 and BBG inhibited BT325 cell growth. The maximum of cell proliferation inhibition rate of GM3 and BBG were 60.28% and 19.33%, respectively. GM3 and BBG were added to medium which has different concentration of EGF. Both GM3 and BBG inhibited the EGF-stimulated BT325 cell growth and the effects of GM3 is far stronger than BBG.

Abstract:By using DNA electrophoresis, flow cytometry (FCM) analysis and laser confocal microscopy (LCM), the change of cell cycle and cytoskeleton of CNE-2Z cells induced by the inhibitors of protein kinase C (PKC) were observed. Cells were respectively treated by straurospine (ST) and sphingosine (SS) at the concentration of 1×10^-6 mol/L and 4×10^-5 mol/L, and cocultured for 24 hours. Ladders of DNA electrophoresis and hypodiploid peaks were discovered in the two treated groups. Compared with the control groups, the cell cycle percentage of S in treated groups by SS was much increased, but the percentage of G1 was significantly decreased (P＜0.05), and the percentage of G2 in ST treated groups was much increased, but both the percentage of S and G1 were much decreased (P＜0.01). The well-distributed chromatin DNA was seen in untreated cells, but fragmentary DNA in treated cells. The granuliform microfilament regularly lined along and formed the intact cell morphology in untreated group cells. The microflament clustered in disorder and formed clusters in induced cells. The results showed that the cell cycle and cytoskeleton may contribute to the CNE-2Z cells apoptosis induced by the inhibitors of protein kinase C.

Abstract:Four different experssion vectors were constructed by cloning foreign gene which encode Schistosoma japonicum 26K antigen(Sj26GST) into Escherichia coli-Mycobacteria shuttle plasmid pBCG-2000 and their expression efficiency were investigated in Mycobacterium smegmatis. The plasmid which contains promoter of human Mycobacterium tuberculosis heat shock protein 70(hsp70) was digested with NcoⅠ and modified with two different ways to lead to two kinds of SD squences, and then ligated with Sj26GST encoding gene. The DNA fragment contained hsp70 promoter and Sj26GST gene was cloned into pBCG-2000, and finally four recombinant mycobacterial expression vectors that are different in SD sequence, orientation and copy number were selected. The expressed native recombinant Sj26GST(rSj26GST) could be observed on SDS-PAGE about at the molecular weight of 26 ku obviously. Analysis with protein density scanning indicated that the expression efficiency that containing double-copy promoter-foreign gene vector was the highest and the expressed protein was about 1.6 folds than that of others. The cloning direction and SD sequence had no significant effect on expression efficiency.

Abstract:A new amperometic biosensor for hydrogen peroxide based on methylene blue incorporated into Nafion membrane as electron transfer mediator was fabricated. It was found that methylene blue incorporated into Nafion membrane by ion-exchanging could effectively transfer electrons between horseradish peroxidase and glassy carbon electrode. Bio-electrocatalytic reduction of hydrogen peroxide at the biosensor was evaluated with respect to solution pH, temperature, operating potential and influences of ascorbic acid etc. The biosensor response exhibited fine selectivity, high sensitivity and a linear dependence on the analytic concentration range 5×10^－7～2×10^－4 mol/L. Response time was less than 30 s.

Abstract:Instead of the NaI scintillation counter used before, a position sensitive detector was used with the position precision 100 μm and the effective working area 50 mm×10 mm. Good SAXS spectrum of liquid samples that can not be measured before was got. The statistical error was decreased and the total time of experiments was shortened.

Abstract:A non-competitive enzyme-linked immunosorbent assay (ELISA) for LpB∶E was developed. Microtiter plates were used as solid-phase and coated with affinity purified goat antibodies to human apo-B. After incubating the antigen in standards and samples with coated plates, a horseradish peroxidase-labelled goat antibodies to human apo-E were added to the plates to estimate the apo-E with apo-B (LpB∶E) by comparing with the standard carried out simultaneously. 120 samples of fasting human plasma randomly collected were quantitated for LpB∶E and the results obtained were briefly discussed.

Abstract:Using acridine orange as fluorescence probe for DNA in cell, measurements of cellular DNA contents of four cases of human breast tumor were done by the system of Hadamard transform microscopic image analysis and conventional microfluorometry respectively, and the analytical results were compared. The results by the two methods are in agreement and both are concordant with pathological diagnosis. It shows that the new instrument for quantitative cytological analysis-Hadamard transform microscopic image analysis system, can provide precise analytical results as that by microfluorometer. In addition, the instrument has outstanding advantages such as high signal to noise ratio, the capability of analyzing two cells or more simultaneously and subtracting background signal synchronously.

Abstract:The effects of gel or electrode buffer strips using agarose with different electroendosmosis of 0, 0.03, 0.08, 0.20 mr on electrophoresis were studied. It was shown that only agarose without electroendosmosis can be used for isoelectric focusing. The electrod buffer strips with different electroendosmosis did not affect SDS PAGE significantly but they affect Native PAGE in certain extent. The higher the electroendosmosis was, the more difficultly it ran.

Abstract:Using the pulse microprogram edited 2D ¹H-¹⁵N HSQC, HSQC-NOESY, HSQC-TOCSY spectra of the ¹⁵N-labeled protein GAL4(62) were abtained, while solvent suppression was achieved by presaturation during the relaxation delay and by the use of spin-lock purge pulses. The important role of those 2D experiments in ¹H assignments of ¹⁵N-labeled proteins was discussed.

Abstract:The kinetic study of the incorporation of exogenous ganglioside GM3 into the lipid bilayer of rabbit sarcoplasmic reticulum (SR) was carried out by trace extraction of Gls and high performance TLC. The results showed that the amount of incorporation is related to GM3 concentration, incubation temperature and time. The optimum incorporation was observed when 8 μmol/L GM3 was incubated with SR vesicles for 90 min at 35℃. The obtained results suggested that the effect of exogenous ganglioside GM3 on SR membrane is not only a simple aqueous phase reaction, but also a kinetic course of the incorporation into the lipid bilayer of SR. Furthermore, the incorporation of GM3 can markedly increase the activity of SR Ca²⁺-ATPase. This provides basic experimental data to study the role of gangliosides in intracellular membranes.

Abstract:A method of improving the recombination efficiency of larger plasmid DNA is introduced. The major manipulation of this method is that the ligation reaction between insertor and vector is separated from cycle reaction of ligation, and they carry out in different reaction condition. Using this method, the efficiency of the recombination between 7.65 kb vector and 1.47 kb insertor is improved obviously. It is a good method for recombination of larger plasmid DNA.

Abstract:The dodecapeptide which is located in the hydrophilic loop connecting helix Ⅳ and Ⅴ of polypeptide D1 of spinach, was synthesized and convalently linked to BSA. Then the complex was injected into a rabbit. Nine weeks later, the specific antiserum was obtained, which showed high titer (1∶256) in immunodiffusion. Western blotting showed that there was an immunoaffinity reaction between the antibody and D1 band of PSⅡ reaction center D1-D2-Cyt b559 complex from spinach chloroplasts. Therefore, this antibody can be used as a probe to detect D1 protein and its degraded fragment during photoinhibition.

Abstract:BIA Technology has been used to study the mechanism of signal transduction in E.coli and the formation of transduction complex. BIA also been used wildly in molecular biology research to study the interaction between transcription factor and its operon (DNA).