Expression, purification and polyclonal antibody preparation of GST-Ccd1 fusion protein / 中国应用生理学杂志

<p><b>AIM</b>By using of Escherichia coli DH5alpha to express GST-Ccd1 fusion protein and which was purified by affinity chromatographic separation. The purified target protein was used to immunize rabbits to prepare polyclonal antibody.</p><p><b>METHODS</b>The previously constructed recombinant prokaryotic expression vector pGEX-5X-1-Ccd1 was transformed into Escherichia coli DH5alpha and was induced to expression by IPTG. The recombinant target protein was expressed with soluble state in Escherichia coli expression system which was separated and purified by chromatographic column stuffed with glutathione Sepharose 4B. The prepared antigen was used to immunize rabbits to get anti-Ccd1 specific rabbit original polyclonal antibody.</p><p><b>RESULTS</b>ELISA data demonstrated that the antibody titer of the serum was up to 1:40 000. Immunohistochemistry analysis indicated that the "home-made" antibody had a specific interaction with Ccd1 protein and which could be used for extended experimental research.</p><p><b>CONCLUSION</b>The anti-Ccd1 polyclonal antibody we had prepared had a high quality of potency and specificity. The antibody was demonstrated by experiments that which could totally fulfill the requirement of immunoblotting and immunohistochemistry study of Ccd1. The antibody provided an useful experimental tool to profoundly research the tissue expression profile, intercellular location and biological function of Ccd1.</p>

Identification of novel genes related to glioma by oligonucleotide microarray / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To identify the genes differentially expressed in development of human glioma, and to study the expression of some genes in different grade gliomas.</p><p><b>METHODS</b>Oligonucleotide microarray (including 218 genes related to neural system development) was adopted and hybridized with probes which were prepared from the total RNAs of glioma specimens and normal brain tissues. Differentially expressed genes between the normal tissues and glioma tissues were assayed after scanning oligonuceltide microarray with ScanArray 4000, and some of these genes such as smad1, Hmp19 and TRIP3 were verified by real-time quantitative PCR(real-time-Q-PCR) method.</p><p><b>RESULTS</b>In comparison with the genes in the normal brain tissue, 5 down-regulated and 5 up-regulated genes in glioma specimens were revealed by means of microarrays, and the expression of smad1, Hmp19 and TRIP3 were verified by real-time-Q-PCR assay.</p><p><b>CONCLUSION</b>Multiple genes play important roles in development of glioma. cDNA microarray technology is a powerful technique in screening for differentially expressed genes between glioma tissues and normal brain tissues. This study is helpful for judgement of invasion and prognosis of gliomas, and provides more target genes for targeted therapy.</p>

Construction of Recombinant Adenovirus Expression Vector of Human Sema4C Gene and Its Expression in Mouse Myoblasts Cell Line C2C12 / 中国生物工程杂志

To generate recombinant adenovirus expression vector of human Sema4C gene and observe its expression in mouse myoblasts cell line C2C12 for ensuring easy access to investigate the role of Sema4C gene during myogenesis. The recombinant plasmid was packaged and amplified after being transfected in HEK293 cells through Lipofectamine. After infecting C2C12 myoblasts with recombinant adenovirus vector, the adenoviral infection efficiency was determined by confocal microscope which showed that the expression of green fluorescence could be detected at 12h and then reached peak at 24h after recombinant adenovirus infection. The infection efficiency was almost 100% confirmed by FACS examination. Detection of WB indicated that the expression of Sema4C in C2C12 of recombinant adenoviral infection group was significantly higher than that of the control group (P

Heat injury induced apoptosis in rat striatum neurons / 中国应用生理学杂志

<p><b>AIM</b>To investigate the effect of hyperthermia on apoptosis of cultivated striatum neurons in the rat.</p><p><b>METHODS</b>After 30 min hyperthermia in 43 degrees, the Ca2+ concentration, the mitochondria membrane potential of the neuron were detected by Laser Scanning Confocal Microscopy (LSCM). The apoptosis of striatum neurons was detected by TUNEL staining.</p><p><b>RESULTS</b>Heat stress at 43 degrees C for 40 min caused an increase in the Ca2+ concentration of striatum neurons and a decrease in the mitochondria membrane potential of the striatum neurons.</p><p><b>CONCLUSION</b>The striatum shows more apoptosis neurons after heat stress.</p>