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1.
FIG. 6.

FIG. 6. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Transduction efficiency of Xrcc4−/− CHO cells infected with an HIV-1 vector. CHO-K1 and XR-1 cells were distributed in 60-mm-diameter dishes at a density of 105 cells per dish. Cells were infected 24 h later with the HIV-1 lacZ vector and scored 2 days postinfection as described in the legend to Fig. 1. The MOI at the 10−2 dilution was 0.0025. The average count of two dishes for each point and standard deviation (error bar) are shown.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
2.
FIG. 3.

FIG. 3. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Effect of reintroduction of DNA-PKCS into the scid background on transduction efficiency by an HIV-1 vector. ST.SCID and 100E cells were distributed in a 96-well plate at a density of 104 cells per well. Cells were infected 24 h later with undiluted virus (MOI of ≈0.01) or the indicated dilutions of the HIV-1 lacZ vector preparations as described in the legend to Fig. 1. Two days postinfection, cells were stained using a β-galactosidase assay (Stratagene), blue cells were counted (two wells/datum point), and digital micrographs were taken of cells infected with the undiluted virus. ST.SCID cells (black columns) and 100E cells (shaded columns) are shown.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
3.
FIG. 4.

FIG. 4. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Transduction efficiencies of normal and scid MEFs infected with a HIV-1 vector encoding an EGFP reporter. Normal and scid MEFs (11th passage) were distributed in 60-mm-diameter dishes at a density of 105 cells per dish. Cells were infected 24 h later with two different dilutions of the HIV-1 EGFP vector (MOI for undiluted cells of ≈0.05) as described in the legend to Fig. 1. Twelve days postinfection, cells were harvested and analyzed by flow cytometry to determine the percentage of EGFP-expressing cells in each sample. The average count of two dishes for each point is shown.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
4.
FIG. 2.

FIG. 2. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Effect of cell density on the efficiency of transduction of MEFs by an HIV-1 vector. Normal MEFs (grey columns) and scid MEFs (black columns) (11th passage) were distributed in a 96-well plate at a density of 5 × 103 to 5 × 104 cells per well, two wells for each point. Cells were infected 24 h later with the HIV-1-lacZ vector at a MOI of ≈0.04 and scored 2 days later as described in the legend to Fig. 1. Mean numbers and standard deviations (error bars) are shown. ex, exponentially growing; sc, subconfluent; c, confluent.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
5.
FIG. 5.

FIG. 5. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Transduction efficiency of scid and normal MEFs infected with an ASV vector. Normal (wild-type) and scid MEFs were distributed into the wells of 24-well plates at 2 × 104 cells per well. After overnight culturing, the cells were infected with undiluted (MOI of ≈0.2) or 1:5 diluted filtered stock of the ASV vector. Forty-eight hours postinfection, the cells were fixed and stained for alkaline phosphatase activity. (Top) Phase-contrast images of the alkaline phosphatase-stained fibroblasts from the cultures infected with the 1:5 dilution of virus. (Bottom) Stained cells were counted after overnight development, and the numbers per well were plotted. Each dilution was assayed in triplicate. The average number of transduced cells per well and standard deviation (error bar) are shown.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
6.
FIG. 7.

FIG. 7. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Killing of ligase IV-deficient cells is integrase dependent after infection under standard conditions but not after spinoculation. Nalm-6 LIG4−/− cells were infected at a MOI of ≈2 (titer determined by transduction of HeLa cells) with the HIV-1 lacZ vectors containing either wild-type integrase or integrase with the inactivating E152A substitution. Cells were either infected under standard conditions, as described in Materials and Methods, or by spinoculation (23). In spinoculation, the vector-cell mixtures were subjected to centrifugation at 1,000 × g for 2 h. (A) Viability was determined by trypan blue exclusion. (B) The relative number of viable cells was determined by counting and expressing the value as a percentage of the cells counted in the mock-infected or mock-spinoculated cultures. Results from standard infection methods(left panels) and with spinoculation (right panels) are shown. Two samples were analyzed for each datum point. Symbols: open circles, mock-infected cells; light grey squares, cells infected with wild-type integrase-containing vector; black triangles, cells infected with the E152A integrase-containing vector.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.
7.
FIG. 1.

FIG. 1. From: Evidence that Stable Retroviral Transduction and Cell Survival following DNA Integration Depend on Components of the Nonhomologous End Joining Repair Pathway.

Transduction efficiencies of normal and scid MEFs infected with an HIV-1 vector. Normal (wild-type) and scid MEFs (11th passage) were distributed in a 96-well plate at a density of 104 cells per well. Cells were infected 24 h later with the HIV-1 lacZ vector (22) for 2 h in the presence of 10 μg of DEAE dextran per ml. Two days postinfection, cells were stained using a β-galactosidase assay kit (Stratagene), and digital micrographs were taken at magnifications of ×4 (top micrographs) and ×20 (bottom micrographs). Results are expressed as the percentage of blue cells in scid cultures compared to normal cultures, with the averages and standard deviations (error bars) obtained from six experiments in which cells were stained and counted 2 to 7 days postinfection. The micrograph shows results with cells infected at a MOI of ≈0.1; averaged data in the bar graph are from experiments with MOIs from 0.0001 to 0.1. The growth curves for scid and normal MEFs are shown at the bottom. Cells were plated at a density of 105 cells per 600-mm-diameter dish, and two plates were counted for each datum point. The average values and standard deviations (error bars) are shown.

René Daniel, et al. J Virol. 2004 August;78(16):8573-8581.

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