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Search results for the GEO ID: GSE30296

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GSM750898

GPL570

Stable expression of BRCA1 in SKOV3 cells - Repeat 1

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750898
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750898/suppl/GSM750898_BRCA1_Clone19_d1.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750898/suppl/GSM750898_BRCA1_Clone19_d1.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

GSM750931

GPL570

Stable expression of BRCA1 in SKOV3 cells - Repeat 2

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750931
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750931/suppl/GSM750931_BRCA1_Clone19_d2.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750931/suppl/GSM750931_BRCA1_Clone19_d2.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

GSM750932

GPL570

Stable expression of BRCA1 in SKOV3 cells - Repeat 3

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750932
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750932/suppl/GSM750932_BRCA1_Clone19_d3.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750932/suppl/GSM750932_BRCA1_Clone19_d3.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

GSM750933

GPL570

Neo Clones of SKOV3 ovarian carcinoma cells - Repeat 1

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750933
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750933/suppl/GSM750933_CONTROL_pc_DNA3_d1.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750933/suppl/GSM750933_CONTROL_pc_DNA3_d1.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

GSM750934

GPL570

Neo Clones of SKOV3 ovarian carcinoma cells - Repeat 2

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750934
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750934/suppl/GSM750934_CONTROL_pc_DNA3_d2.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750934/suppl/GSM750934_CONTROL_pc_DNA3_d2.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

GSM750935

GPL570

Neo Clones of SKOV3 ovarian carcinoma cells - Repeat 3

Mamalian Cultured cells (ovarian carcinoma)

tissue type: ovarian tumor disease type: ovarian carcinoma cell line: SKOV3

Sample_geo_accession	GSM750935
Sample_status	Public on Jul 02 2011
Sample_submission_date	Jun 29 2011
Sample_last_update_date	Jul 02 2011
Sample_type	RNA
Sample_channel_count	1
Sample_organism_ch1	Homo sapiens
Sample_taxid_ch1	9606
Sample_molecule_ch1	total RNA
Sample_extract_protocol_ch1	PROTOCOL:
Sample_extract_protocol_ch1	1. Harvest cells or use cells that are already pelleted before.
Sample_extract_protocol_ch1	Cells grown in suspension or monolayer (1 x 107 cells)
Sample_extract_protocol_ch1	Pellet the cells for 5 min at 300 x g in an RNase-free glass or polypropylene centrifuge tube. Carefully remove all supernatant by aspiration. Use tripsin in case of monolayer. Cells can also be lysed directly on the plate using desired amount of lysis buffer.
Sample_extract_protocol_ch1	Note: Incomplete removal of the cell-culture medium will inhibit lysis and dilute the lysate, affecting the conditions for binding of RNA to the RNeasy membrane.
Sample_extract_protocol_ch1	2. Disrupt cells by addition of Buffer RLT.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	For pelleted cells, loosen the cell pellet thoroughly by flicking the tube. Add the appropriate volume of Buffer RLT according the following table and proceed to step 3.
Sample_extract_protocol_ch1	RNeasy column Number of cells Buffer RLT (μl)
Sample_extract_protocol_ch1	Mini < 5 x 106 350
Sample_extract_protocol_ch1	Mini 5 x 106 – 1 x 107 600
Sample_extract_protocol_ch1	Note: Ensure that β-ME is added to Buffer RLT before use
Sample_extract_protocol_ch1	3. Homogenize cells using homogenizer for at least 30 seconds. Alternatively, vortex the sample for 10 s, and pass the lysate at least 5–10 times through an 18–20-gauge needle fitted to an RNase-free syringe.
Sample_extract_protocol_ch1	Note: Incomplete homogenization will lead to significantly reduced yields and can cause clogging of the RNeasy column. Homogenization with homogenizers generally results in higher total RNA yields than with other homogenization methods.
Sample_extract_protocol_ch1	4. Add 1 volume (350 μl or 600 μl) of 70% ethanol to the homogenized lysate, and mix thoroughly by shaking vigorously. Do not centrifuge.
Sample_extract_protocol_ch1	5. Apply the total sample (up to 700 μl), including any precipitate that may have formed, to an RNeasy mini column placed in a 2 ml collection centrifuge tube. Close the tube gently, and centrifuge for 30-45 second at > 8000 g (> 10000 rpm). Discard the flow-through. If the volume exceeds 700 μl load the column again with the remaining extract solution.
Sample_extract_protocol_ch1	6. Add 700 μl Buffer RW1 to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through and the collection tube.
Sample_extract_protocol_ch1	7. Transfer the column to a fresh 2 ml collection tube Add 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 30-45 seconds at > 8000 g (> 10000 rpm) to wash the column. Discard the flow-through.
Sample_extract_protocol_ch1	Note: Buffer RPE is supplied as a concentrate. Ensure that ethanol is added to Buffer RPE before use.
Sample_extract_protocol_ch1	8. Add another 500 μl Buffer RPE to the RNeasy column. Close the centrifuge tube gently, and centrifuge for 5 min at > 8000 g (> 10000 rpm) to dry the RNeasy silica-gel membrane. Transfer the columns in a separate tube and spin it for 1 minute at high speed.
Sample_extract_protocol_ch1	It is important to dry the RNeasy membrane since residual ethanol may interfere with downstream reactions. This centrifugation ensures that no ethanol is carried over during elution.
Sample_extract_protocol_ch1	Prepared by Tapas Saha
Sample_extract_protocol_ch1	9. To elute, transfer the RNeasy column to a new 1.5 ml eppendorf tube. Pipet 30-50 μl of RNase-free water directly onto the RNeasy silica-gel membrane. Close the tube gently. Let it stand for 1 min, and then centrifuge for 1 min at > 8000 g (> 10000 rpm).
Sample_extract_protocol_ch1	10. Repeat the elution step (step 9) as described with a second volume of RNase-free water. To obtain a higher total RNA concentration, this second elution step may be performed by using the first eluate.
Sample_extract_protocol_ch1	11. Load a Formamide/Denatured RNA gel (Different protocol) to visualize the quality of the RNA isolated. At the same time measure the concentration of the RNA by spectrophotometer.
Sample_label_ch1	biotin
Sample_label_protocol_ch1	One-Cycle Target Labeling Assay
Sample_hyb_protocol	Cenechip Eukaryotic Target Hybridization protocol
Sample_scan_protocol	Genechip expression washing, stain and scan user manual,P/N 702731
Sample_data_processing	RMA using R/Bioconductor
Sample_platform_id	GPL570
Sample_contact_name	Xin,James,Li
Sample_contact_institute	Georgetown University
Sample_contact_address	4000 Reservoir Rd Suite 180
Sample_contact_city	Washington
Sample_contact_state	DC
Sample_contact_zip/postal_code	20007
Sample_contact_country	USA
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750935/suppl/GSM750935_CONTROL_pc_DNA3_d3.CEL.gz
Sample_supplementary_file	ftp://ftp.ncbi.nlm.nih.gov/geo/samples/GSM750nnn/GSM750935/suppl/GSM750935_CONTROL_pc_DNA3_d3.CHP.gz
Sample_series_id	GSE30296
Sample_data_row_count	54675

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