Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT01586 (G418)
 2,237 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A recombinant bovine leukemia virus (BLV) was constructed in which the X region was replaced with the bacterial neomycin resistance gene controlled by the simian virus 40 early promoter. This virus, termed BLV-SVNEO, is a self-packaging, activator-dependent retroviral vector. Introduction of the plasmid pBLV-SVNEO into mammalian cells resulted in constitutive expression of the neo gene, whereas the BLV structural genes, gag, pol, and env, were expressed only in the presence of the two regulatory proteins, Tax and Rex. The production and release of recombinant virus by cells transfected with pBLV-SVNEO were proportional to the number of G418-resistant colonies that developed after susceptible cells were exposed to the filtered culture medium. BLV-SVNEO was able to infect cell lines of human, bovine, canine, feline, and murine origin. BLV-producing cell lines were resistant to superinfection with BLV-SVNEO. This cell-virus system should facilitate molecular genetic studies of BLV and will provide a rapid, quantitative measure of BLV infectivity in a variety of cell types. These studies also demonstrate the feasibility of using activator-dependent retroviral vectors such as BLV-SVNEO to deliver foreign genes into cells and eventually animals.
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PMID:Construction of a recombinant bovine leukemia virus vector for analysis of virus infectivity. 168 85

Recently, amniotic fluid was suggested as a new source for stem-cell research and tissue engineering approaches. In order to enable isolation of stem cells and establishment of lines of such cells with an undifferentiated phenotype we have introduced green fluorescent protein regulated by the promoters of the stem cell-specific genes, Oct-4 or Rex-1, into human amniotic fluid cells. For the introduction of DNA into human amniotic fluid cells, we have optimized a specific transfection protocol. We found that human amniotic fluid contains cell populations which are able to activate these promoters. These undifferentiated cells expressing green fluorescent protein can be analysed on a flow cytometer. In addition, we have introduced a plasmid harboring a neomycin-resistance gene under the control of the Oct-4 promoter. G418 selection allowed the isolation of undifferentiated stem cells expressing Oct-4 protein out of human amniotic fluid samples. Our findings confirm the existence of stem cells within amniotic fluid. In addition, the ability to transfect human amniotic fluid cells and to isolate stem-cell marker-positive cells will provide the means to study and manipulate these cells for the purpose of basic and applied research.
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PMID:Activation of ectopic Oct-4 and Rex-1 promoters in human amniotic fluid cells. 1627 76

The pluripotency of mouse embryonic stem (ES) cells is maintained by self-renewal. To screen for genes essential for this process, we constructed an RNA interference (RNAi) library by inserting subtracted ES cell cDNA fragments into plasmid containing two opposing cytomegalovirus promoters. ES cells were transfected with individual RNAi plasmids and levels of the pluripotency marker Oct-4 were monitored 48 hours later by real time RT-PCR. Of the first 89 RNAi plasmids characterized, 12 downregulated Oct-4 expression to less than 50% of the normal level and 7 of them upregulated Oct-4 expression to more than 150% of the normal level. To investigate their long-term effect on self-renewal, ES cells were transfected by these 19 RNAi plasmids individually and G418-resistant colonies were subjected to alkaline phosphatase (AP) staining after 7 days selection. Except for 4 plasmids that caused cell death, the ratio of AP positive colonies was repressed to less than 60% of the control group by the other 15 plasmids and even below 20% by 10 plasmids. The cDNA fragments in these 10 plasmids correspond to eight genes, including Zfp42/Rex-1, which was chosen for further functional analysis. RNAi knockdown of Zfp42 induced ES cells differentiate to endoderm and mesoderm lineages, and overexpression of Zfp42 also caused ES cells to lose the capacity of self-renewal. Our results indicate that RNAi screen is a feasible and efficient approach to identify genes involved in ES cells self-renewal. Further functional characterization of these genes will promote our understanding of the complex regulatory networks in ES cells.
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PMID:Screening for genes essential for mouse embryonic stem cell self-renewal using a subtractive RNA interference library. 1696 Jan 29