Human BBC3 (PUMA) knockout HeLa cell line
- Advanced Validation
- What is this?
Be the first to review this product! Submit a review
|
(0 Publication)
BBC3 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, 10 bp deletion in exon 2 and 1 bp insertion in exon 2 and 22 bp deletion in exon 2. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
View Alternative Names
BBC3_HUMAN, Bcl-2-binding component 3, JFY-1, PUMA alpha, PUMA/JFY1, p53 up-regulated modulator of apoptosis
- WB
Lab
Western blot - Human BBC3 (PUMA) knockout HeLa cell line (AB261832)
Lanes 1 - 4 : Merged signal (red and green). Green - anti-Puma antibody observed at 25 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.
anti-Puma antibody was shown to react with Puma in wild-type HeLa cells in Western blot with loss of signal observed in BBC3 knockout cell line ab261832 (BBC3 knockout cell lysate ab256848). Wild-type HeLa and BBC3 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with anti-Puma antibody and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
All lanes:
Anti-Puma antibody at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
BBC3 knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human BBC3 (PUMA) knockout HeLa cell line (ab261832)
Lane 3:
A549 cell lysate at 20 µg
Lane 4:
PC-3 cell lysate at 20 µg
false
- Sanger seq
Unknown
Sanger Sequencing - Human BBC3 (PUMA) knockout HeLa cell line (AB261832)
Allele-1 : 22 bp deletion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human BBC3 (PUMA) knockout HeLa cell line (AB261832)
Allele-2 : 10 bp deletion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human BBC3 (PUMA) knockout HeLa cell line (AB261832)
Allele-3 : 1 bp insertion in exon 2.
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
Handling procedures
Initial handling guidelines
Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.
Subculture guidelines
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
Culture medium
DMEM (High Glucose) + 10% FBS
Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
PUMA functions as a pivotal mediator of apoptosis in response to diverse stimuli such as DNA damage and oncogenic stress. As part of the Bcl-2 family PUMA does not typically form a complex with other proteins but functions directly by binding and neutralizing anti-apoptotic Bcl-2 family proteins. This action results in the release of caspase-activating factors from mitochondria essential for programmed cell death.
Pathways
Several cellular pathways incorporate PUMA to regulate apoptosis. PUMA is notably included in the p53 pathway where p53 transcriptionally activates PUMA following cellular stress or DNA damage. The presence of related proteins like Bax and Bim in the mitochondrial apoptotic pathway links these signals to the mitochondrial death machinery facilitating apoptosis.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Complementary Products
Cell Lines & Lysates
AB265069
Human FOXO3 (FOXO3A) knockout HEK-293T cell line
Advanced Validation
- 0
- 0
Cell Lines & Lysates
AB261873
Human HIST1H1C (Histone H1.2) knockout A549 cell line
Advanced Validation
- 0
- 0
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com