Expression of wnt5a gene in hematologic diseases and leukemic cell lines / 中国实验血液学杂志

This study was aimed to investigate the expression level of Wnt5a gene in some hematologic diseases and leukemic cell lines so as to provide a basis for further research of Wnt5a role and its mechanism in hematologic malignancies. The mononuclear cells of peripheral blood and bone marrow were isolated by human lymphocytic isolation solution. The expression of Wnt5a gene in specimen of 31 cases and three leukemic cell lines (Jurkat, K562, HL-60) were detected by RT-PCR. The results showed that in four out of five AML cases, negative or weak positive expressions were observed and negative expressions were observed also in K562 and HL-60 cells. Only in one AML case with complete remission and Jurkat cells the strong positive expressions were observed. The negative expression was observed in all six CML cases. In three out of four ALL cases, the expression was positive or weak positive and one negative. The expressions in two CLL cases were negative. Out of two MM cases, the expression in one was weak positive and in other was negative. Out of three lymphoma cases, the expression in one case was weak positive and in other two cases were negative. There were positive or weak positive expressions in two cases of AA, two cases of IDA, three cases of ITP, one cases of PV and ET cases. It is concluded that there have obvious down-regulated or lost expression of Wnt5a gene in 31 cases of hematologic disease and myelocytic leukemic cell lines except ALL samples. Nevertheless there have general positive expression of Wnt5a in cases of non-malignant hematologic diseases. These results suggest that the genesis of myelocytic leukemia is related to the down-regulated expression of Wnt5a.

Expression of multi-copy and immuno-reactivity of recombinant type-specific epitope of herpes simplex virus type 1 / 中华男科学杂志

<p><b>OBJECTIVE</b>To express a multi-copy specific epitope recombinant protein of herpes simplex virus type 1 (HSV-1) with immuno-reactivity.</p><p><b>METHODS</b>Multi-copy genes with a specific epitope of HSV-1-glycoprotein G 112-127 were constructed by DNA recombination and cloned in E. coli JM109 pGEM-5Zf. The positive recombinants were determined by SDS-PAGE and Western blotting.</p><p><b>RESULTS</b>The recombinants with 4, 8, 16 and 32 copies of gG 112-127 were obtained. The 8-copy recombinant was expressed by 17.5%, mainly as inclusion body. And it reacted with antiserum HSV-1, but not with antiserum HSV-2.</p><p><b>CONCLUSION</b>The HSV-1-gG112-127 recombinant could be used to distinguish HSV-1 and HSV-2 in ELISA.</p>

Advances in analysis techniques of phosphoproteome / 生物工程学报

In eukaryotes protein phosphorytion is a key event. By reversible protein phosphorylation eukaryotes control many cellular processes including signal transduction, gene expression, the cell cycle etc. Phosphoproteomics involves identification of phosphoproteins and phosphopeptides, localization of the exact residues that are phosphorylated and quantitation of phosphorylation. Because protein phosphorylation is a dynamic process, and it is present at low abundance within cells, and the phosphorylated sites on proteins might vary, and mass spectrometry (MS) signals from phosphopeptides are usually suppressed etc., so phosphoprotein analysis have more difficulties than nonphosphoprotein. In this article, we outline several analysis techniques for separation, identification and quantitation of phosphorylated proteins and peptides, and discuss the progress in these techniques. At present, MS is still an essential core identification technology for phosphoproteomic studies, To search better enrichment strategies are the main challenges in this rapidly evolving field. A major goal of quantitative proteomics is precise quantification and identification of proteins in complex mixtures. A common method for quantitative proteome analysis is the stable isotope labeling method. Today there is no single method that supersedes all others techniques for Phosphoproteomic studies. With continued development of sample preparation techniques and instrumentation, it should be possible to perform a global analysis of protein phosphorylation.

Expression of heat-labile enterotoxin and the strategy of purification and storage / 生物工程学报

Heat-labile enterotoxin (LT) from Escherichia coli is a bacterial protein toxin with an AB5 hexamer structure. LT is a powerful mucosal adjuvant when co-administered with soluble antigens. However, its use in mucosal immunity is inconvenient because of its low yield and depolymerization during long-term storage under normal condition. In this study, we report an efficient expression system and optimized purification and storage strategy of LT. A gene encoding LT was cloned into the vector pET11c and transformed in E. coli BL21(DE3). By growing this strain on modified M9-CAA medium, LT was expressed efficiently. About 46mg/L LT could be purified from the supernatant of bacteria lysate. Using D(+)-Immobilized galactose column, LT could be purified at a wide pH range with various elution buffers. The optimized elution buffers are TEAN (pH 7.3) containing 0.3mol/L galactose and carbonate buffer (pH 10.4) containing 0.3mol/L galactose. After dried by freeze and placed in 4 degrees C, LT dissolved in TEAN (pH 7.3) and carbonate buffer (pH 10.4) were assayed by HPLC. The results indicated that the integrity of AB5 hexamer was kept well. LT could undergo long-term storage under this condition. This was proved to be an optimized strategy of LT storage. The results of GM1 binding assay and toxicity assay showed that the purified recombinant LT has normal biological character.

STUDIES ON LIQUID STATE CULTURE CONDITIONS OF HELICOBACTER PYLORI / 微生物学通报

There are still many problems at liquid state culture especially large-scale culture for Helicobacter pylori at present. Using analytical technique of single factor test statistics, a suitable medium and shaking culture conditions for Helicobacter pylori have been selected. The experimental results showed: under the optimal shaking culture conditions Hp could be growed well enough in Brucella broth with ?-CD. The thalli weight reached 0.6g/100mL.