[Therapeutic potential of the neural stem cells against ischemic brain injury].
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Animals↗
Biomedical subjects
Ming Fan
Publications and source records attributed to Ming Fan.
MProbe: computer aided probe design for oligonucleotide microarrays.
The present work describes a complete probe design software system for oligonucleotide microarrays based on Kane's research on probe sensitivity and specificity (Kane's rule). Combining Kane's rule and traditional criteria for probe design we constructed MProbe, the software system for oligonucleotide microarrays using Java. The general criteria for probe design are: (1) probes may have different lengths that range from 20 to 100 bases; (2) they should have a similar melting temperature (Tm) or GC content; (3) they should not contain stable secondary structures; and (4) they abide by Kane's rule.
Computational Biology↗
Construction and Characterization of Recombinant Adenoviruses Expressing Biologically Active Human Brain-derived Neurotrophic Factor and Neurotrophin-3.
In order to deliver brain-derived neurotrophic factor(BDNF) and neurotrophin-3(NT3) into CNS to prevent or reduce degeneration of CNS neurons in human neurodegenerative diseases and neuron injuries, recombinant adenoviruses Ad-BDNF and Ad-NT3 were constructed. BDNF and NT3 genes were inserted into an E1-substituted adenovirus shuttle plasmid, respectively, and were driven by the human cytomegalovirus immediate-early gene promoter/enhancer. After cotransfection into 293 cells with the adenovirus plasmid pJM17, the recombinant adenovirus Ad-BDNF and Ad-NT3 were propagated in 293 cells via homologous recombination. After two rounds of CsCl centrifugation, the human recombinant adenovirus Ad-BDNF and Ad-NT3 were obtained with titer of 1x10(12) and 8x10(11) pfu/ml, respectively. To examine the expression of BDNF and NT3, HeLa cells were infected with Ad-BDNF and Ad-NT3, respectively. RT-PCR, Western blot and ELISA analysis results showed that BDNF and NT3 genes could be transcribed and translated in Ad-BDNF and Ad-NT3 infected HeLa cells. And the culture media containing 10% conditioned medium of Ad-BDNF and Ad-NT3 infected Hela cells could induce the neurite outgrowth from E8 dorsal root ganglion neurons.
Journal Article↗
Eukaryotic Expression in COS7 Cells and Biological Activity of Human Brain-derived Neurotrophic Factor (BDNF).
In order to study the expression of human brain-derived neurotrophic factor in eukaryotic cells, hBDNF cDNA was cut off from MI3mpl8-hBDNF plasmid and inserted into the eukaryotic expression vector pCMV4. Using lipofectin methods, the recombinant expression plasmid pCMV4-hBDNF was transfected into COS7 cells for transient expression. RNA slot hybridization analysis and immunocytoreaction analysis demonstrated that the BDNF gene can be transcripted and translated in COS7 cells. The human rBDNF secreted by transfected COS7 cells can promote the development and survival of dopaminergic neurons.
Journal Article↗
RNA aptamers specific for bovine thrombin.
Bovine thrombin is widely used in clinical wound healing after surgery. There is 85% homology between bovine thrombin and human thrombin, so most antibodies against bovine thrombin cross-react with human thrombin. Rare antibodies against bovine thrombin but not cross-reacting with human thrombin have been reported. RNA ligands (aptamers) have been used to bind to target molecules with sometimes higher specificity than antibodies. Here we report the isolation of aptamers specific for bovine thrombin by systematic evolution of ligands by exponential enrichment (SELEX) from an RNA pool containing a 25-nucleotide randomized region. After seven rounds of selection, two aptamers specific for bovine thrombin were identified with a K(d) of 164 and 240 nM, respectively. Significantly, these aptamers do not bind to human thrombin. Secondary structure prediction revealed potential stem-loop structures for these RNAs. Both RNA aptamers inhibit only bovine thrombin-catalyzed fibrin clot formation in vitro. Competition assay results suggested that the RNA aptamers might bind to the electropositive domain of bovine thrombin, that is, heparin-binding site, instead of fibrinogen-recognition exosite. The resulting bovine-specific thrombin inhibitor might be used in some clinical applications when bovine thrombin activity needs to be contained or in research where human and bovine thrombin need to be distinguished.
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