2010
DOI: 10.1089/hum.2009.107
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Robust, Reversible Gene Knockdown Using a Single Lentiviral Short Hairpin RNA Vector

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Cited by 29 publications
(28 citation statements)
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“…To generate ReNcells VM with doxycycline-inducible knock-down of USP9X, cells were transduced with a lentiviral vector 43 containing either a “scrambled” shRNA sequence: 5′ACTACCGTTGTTATAGGTGTTCAAGAGACACCTA TAACAACGGTAGT3′ with no homology to any known gene, or shRNA targeting two independent USP9X sequences “2193” 5′GCTTGATCCTTCCCTGTTAAC3′ (the first base pair is 2193 in the human USP9X sequence) or “4774” 5′GCCATAGAAGGCACAGGTAGT3′. Successfully transduced pools of ReNcells VM were isolated by FACS based on constitutive EGFP expression 43 . These transduced pools were used in subsequent experiments.…”
Section: Methodsmentioning
confidence: 99%
“…To generate ReNcells VM with doxycycline-inducible knock-down of USP9X, cells were transduced with a lentiviral vector 43 containing either a “scrambled” shRNA sequence: 5′ACTACCGTTGTTATAGGTGTTCAAGAGACACCTA TAACAACGGTAGT3′ with no homology to any known gene, or shRNA targeting two independent USP9X sequences “2193” 5′GCTTGATCCTTCCCTGTTAAC3′ (the first base pair is 2193 in the human USP9X sequence) or “4774” 5′GCCATAGAAGGCACAGGTAGT3′. Successfully transduced pools of ReNcells VM were isolated by FACS based on constitutive EGFP expression 43 . These transduced pools were used in subsequent experiments.…”
Section: Methodsmentioning
confidence: 99%
“…Cells were cultured in 3D Matrigel cultures (BD Biosciences) and stained with immunofluorescently tagged antibodies, as described previously (17). A Zeiss LSM 510 confocal microscope was used to EF-1a promoter (67). Gateway technology (Invitrogen) was used for the transfer of miR-34a into the pLV711G vector to create a single lentiviral vector enabling doxycycline-responsive expression of miR-34a.…”
Section: Methodsmentioning
confidence: 99%
“…HCT-116 O and HCT-116 N cells were also engineered to stably express histone H2B-green fluorescent protein (H2B-GFP). Histone H2B-GFP from the pBOS-H2B-GFP vector (Becton Dickinson) was subcloned into the SalI/ NotI restriction sites of the pENTR vector (Life Technologies), and then moved into the Gateway-compatible lentiviral vector pLV418G (36). The construct was packaged into lentivirus which was used to transduce cells.…”
Section: Methodsmentioning
confidence: 99%