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Accession IconSRP075430

Homo sapiens Raw sequence reads

Organism Icon Homo sapiens
Sample Icon 2 Downloadable Samples
Technology Badge IconIllumina HiSeq 2500

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In order to validate our method to develop immortalized cellular models of microglia, we used RNA-seq to confirm the microglial phenotype of a representative clone (C20, a clonal population), which derived from the transformation of human primary microglia (human microglia purchased cryopreserved after purification from Sciencell, Cat. #1900). Transformation was carried out with vesicular stomatitis virus G envelop simian virus 40 large T antigen viral particles (VSVG SV40), containing the pBABE-puro SV40 LT construct (Addgene, Plasmid #13970), followed by VSVG containing the human telomerase reverse transcriptase (hTERT)-neomycin (pBABE-neo-hTERT) construct (Addgene, Plasmid #1774). Immortalized cells were selected in the presence of 2 µg/mL puromycin and 600 µg/mL neomycin. Individual clonal populations, including clone C20, were allowed to grow for approximately 4 weeks prior to further testing. In addition, we also used RNA-seq to verify the capacity of these cellsto respond to an inflammatory stimulus (TNF-a).
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