Search results for the GEO ID: GSE7324  |
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GSM ID | GPL ID |
Select for analysis |
Title |
Source name |
Description |
Characteristics |
GSM176671 | GPL1261 |
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GFP-transduced_rep1
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GFP/MigR1 (vector-only)-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using control (GFP/MigR1) retrovirus. Retroviral (GFP) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176672 | GPL1261 |
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GFP-transduced_rep2
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GFP/MigR1 (vector-only)-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using control (GFP/MigR1) retrovirus. Retroviral (GFP) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176673 | GPL1261 |
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GFP-transduced_rep3
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GFP/MigR1 (vector-only)-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using control (GFP/MigR1) retrovirus. Retroviral (GFP) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176674 | GPL1261 |
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AML1/ETO-transduced_rep1
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AML1/ETO-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using AML1/ETO retrovirus. Retroviral (AML1/ETO) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176675 | GPL1261 |
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AML1/ETO-transduced_rep2
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AML1/ETO-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using AML1/ETO retrovirus. Retroviral (AML1/ETO) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176676 | GPL1261 |
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AML1/ETO-transduced_rep3
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AML1/ETO-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on three samples using AML1/ETO retrovirus. Retroviral (AML1/ETO) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176677 | GPL1261 |
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W692A-transduced_rep1
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W692A-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on four samples using W692A retrovirus. Retroviral (W692A) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176678 | GPL1261 |
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W692A-transduced_rep2
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W692A-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on four samples using W692A retrovirus. Retroviral (W692A) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176679 | GPL1261 |
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W692A-transduced_rep3
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W692A-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on four samples using W692A retrovirus. Retroviral (W692A) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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GSM176680 | GPL1261 |
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W692A-transduced_rep4
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W692A-transduced, live, lineage-negative, Sca1 cKit double-positive bone marrow cells
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C57BL/6, 5-6 weeks old
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Primary bone marrow mononuclear cells were harvested from 5-6 week old C57BL/6 mice (Jackson Labs, Bar Harbor, ME) and subjected to immunomagnetic negative selection using the Lineage cell depletion Kit (Miltenyi Biotech, Auburn, CA) to enrich for stem and progenitor cells (Lin- cells). To further enrich the stem cell compartment, Lin- cells underwent positive selection for the expression of Sca1 (Lin-Sca1+;LS) using the Anti-Sca-1 MicroBead Kit (Miltenyi Biotech). LS cells (6x105 in 3ml) were plated in ultra-low adhesion 6-well plates (Costar, Corning, NY) and incubated overnight at 37oC, 5% CO2 in transplant media [RPMI, 20% FCS, penicillin and streptomycin, 10 ng/ml IL-3, 20 ng/ml IL-6 (R&D Systems, Minneapolis, MN), 10 ng/ml SCF (Stem Cell technologies, Vancouver, Canada)]. Twelve hours later, 6x105 cells in 2ml of fresh transplant media were added to 6-well plates (Cellstar, GBO, NC) coated with 100 mg Retronectin (Takara, Madison WI). Independent retroviral transductions were performed on four samples using W692A retrovirus. Retroviral (W692A) supernatants (2 ml), 4 µl of 40 mg/ml polybrene and 40µl 1M Hepes were added to each well and the cells centrifuged for 90 minutes at 1400 g, 37oC. Cells were subjected to a second round of transduction 24 hours later as described above. 48 hours following the second transduction, cells were collected (kept as independent samples) and stained with PE conjugated anti-cKit (BD Bioscience), APC conjugated anti-Sca1 (Ebioscience), and biotin conjugated anti-lineage cocktail (Miltenyi Biotech). PerCP Cy5.5 conjugated Streptavidin (BD Bioscience) was added to identify differentiated cells marked with biotin conjugated antibodies. 7-AAD (Molecular Probes) was used to exclude dead cells from the sort. 25,000 transduced (GFP+), live, Lin- (7-AAD/PerCP Cy5.5-), Sca1+ cKit+ (APC-PE ++) cells were sorted directly into 600ul of RLT RNA lysis buffer (Qiagen) using a BD FACSAria cell sorter (BD Bioscience).
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