CDKN1B KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9,
Homozygous: 52 bp deletion-3 bp insertion in Exon 1.
Images
Key facts
- Cell type
RAW 264.7
- Species or organism
Mouse
- Tissue
Lymphatic
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
Knockout achieved by using CRISPR/Cas9,
Homozygous: 52 bp deletion-3 bp insertion in Exon 1
Alternative names
AA408329, AI843786, CDKN 1B, CDKN 4, CDN1B_HUMAN, Cdki1b, Cyclin-dependent kinase inhibitor 1B, Cyclin-dependent kinase inhibitor 1B (p27, Kip1), Cyclin-dependent kinase inhibitor p27, Cyclin-dependent kinase inhibitor p27 Kip1, KIP 1, MEN1B, MEN4, OTTHUMP00000195098, OTTHUMP00000195099, P27-like cyclin-dependent kinase inhibitor, p27, p27 Kip1
Recommended products
CDKN1B KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9,
Homozygous: 52 bp deletion-3 bp insertion in Exon 1.
Alternative names
AA408329, AI843786, CDKN 1B, CDKN 4, CDN1B_HUMAN, Cdki1b, Cyclin-dependent kinase inhibitor 1B, Cyclin-dependent kinase inhibitor 1B (p27, Kip1), Cyclin-dependent kinase inhibitor p27, Cyclin-dependent kinase inhibitor p27 Kip1, KIP 1, MEN1B, MEN4, OTTHUMP00000195098, OTTHUMP00000195099, P27-like cyclin-dependent kinase inhibitor, p27, p27 Kip1
Key facts
- Cell type
RAW 264.7
- Species or organism
Mouse
- Tissue
Lymphatic
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
- Mutation description
Knockout achieved by using CRISPR/Cas9,
Homozygous: 52 bp deletion-3 bp insertion in Exon 1- Disease
Carcinoma
- Concentration
Properties
- Gene name
CDKN1B
- Gene editing type
Knockout
- Gene editing method
CRISPR technology
- Knockout validation
Sanger Sequencing, Western blot
- Zygosity
Homozygous
Cell culture
- Biosafety level
EU: 2 US: 2
- Adherent/suspension
Adherent
- Gender
Male
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 for bath 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 all remaining cells into an ultra-low attachment 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.
5. Once confluent passage into an appropriate flask at a density of 2x105 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
- Grow in ultra-low attachment flasks.
- Do not wash or use dissociation reagent. Cells are semi-adherent. Tap flask sharply to remove adhered cells.
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x105 cells/mL is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
- Culture medium
DMEM + 10% FBS
- Cryopreservation medium
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Storage
- Shipped at conditions
Dry Ice
- Appropriate long-term storage conditions
-196°C
Notes
Recommended control: Human wild-type RAW 264.7 cell line (Mouse wild-type RAW 264.7 cell line ab275474)
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.
Supplementary info
- Activity summary
The protein known as p27 KIP1 also referred to as p27 CDKN1B or KIP1 is a member of the kinase inhibitory protein family. It plays an important role in cell cycle regulation by inhibiting cyclin-dependent kinases (CDKs). p27 KIP1 weighs approximately 27 kDa and can be found in various tissues where it regulates cell proliferation. The protein functions as a suppressor by binding to cyclin-CDK complexes preventing the transition from G1 phase to S phase in the cell division cycle.
- Biological function summary
The function of p27 KIP1 involves its role in controlling cell growth and division. It achieves this by becoming a part of larger protein complexes involving CDKs and cyclins. By directly interacting with these complexes p27 KIP1 modulates cell cycle progression therefore acting as a brake on cellular proliferation. The protein is important in maintaining proper cell cycle checkpoints and preventing uncontrolled cell growth which is essential for normal cellular functioning.
- Pathways
The involvement of p27 KIP1 centers on cell cycle regulation and signaling pathways such as the PI3K/AKT pathway. Its interaction with CDKs and cyclins situates it within the core mechanisms that determine cell division timing. p27 KIP1 operates alongside other proteins like cyclin D and CDK4/6 fitting into the regulatory intricacies of these pathways. Proper functioning of these pathways ensures cellular homeostasis and prevents the development of oncogenic processes.
- Associated diseases and disorders
Dysfunction of p27 KIP1 has links to cancer and neurodegenerative diseases. Low levels or mutations can lead to uncontrolled cell proliferation contributing to the development and progression of cancers such as breast cancer. p27 KIP1's role in neurodegenerative diseases involves its regulation of neuronal cell cycle re-entry with abnormalities potentially exacerbating conditions like Alzheimer's disease. In these contexts its interaction with proteins such as cyclin E and CDK2 becomes particularly relevant in understanding disease mechanisms.
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3 product images
Western blot - Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell line (ab281619)
False colour image of Western blot: Anti-p27 KIP 1 antibody [EPFHCR16] staining at 1/1000 dilution shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution shown in red. In Western blot Anti-p27 KIP 1 antibody [EPFHCR16] ab92741 was shown to bind specifically to p27 KIP 1. A band was observed at 28 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CDKN1B knockout cell line ab281619 (knockout cell lysate Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell lysate ab282970). To generate this image wild-type and CDKN1B knockout RAW 264.7 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-p27 KIP 1 antibody [EPFHCR16] (Anti-p27 KIP 1 antibody [EPFHCR16] ab92741) at 1/1000 dilution
Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg
Lane 2: CDKN1B knockout RAW 264.7 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 22 kDa
Observed band size: 28 kDa
Western blot - Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell line (ab281619)
False colour image of Western blot: Anti-p27 KIP 1 antibody [Y236] staining at 1/5000 dilution shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution shown in red. In Western blot Anti-p27 KIP 1 antibody [Y236] ab32034 was shown to bind specifically to p27 KIP 1. A band was observed at 28 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in CDKN1B knockout cell line ab281619 (knockout cell lysate Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell lysate ab282970). To generate this image wild-type and CDKN1B knockout RAW 264.7 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-p27 KIP 1 antibody [Y236] (Anti-p27 KIP 1 antibody [Y236] ab32034) at 1/5000 dilution
Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg
Lane 2: CDKN1B knockout RAW 264.7 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 22 kDa
Observed band size: 28 kDa
Sanger Sequencing - Mouse CDKN1B (p27 KIP 1) knockout RAW 264.7 cell line (ab281619)
52 bp deletion-3 bp insertion in Exon 1
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