CBX4 KO cell line available to order. KO validated by Next Generation Sequencing. Free of charge wild type control provided. Knockout achieved by CRISPR/Cas9; X = 7 bp deletion, 1 bp insertion, 10 bp deletion.
CBX4_HUMAN, Cbx4 chromobox homolog 4 (Drosophila Pc class), Chromobox homolog 4, Chromobox homolog 4 (Pc class homolog, Drosophila), Chromobox protein homolog 4, E3 SUMO-protein ligase CBX4, NBP 16, NS5ATP1 binding protein 16, PC 2, Pc class 2 homolog, Pc class homolog, Pc class homolog Drosophila, Polycomb 2 homolog, hPc 2
CBX4 KO cell line available to order. KO validated by Next Generation Sequencing. Free of charge wild type control provided. Knockout achieved by CRISPR/Cas9; X = 7 bp deletion, 1 bp insertion, 10 bp deletion.
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.
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.
CBX4 also known as Chromobox protein homolog 4 or Polycomb 2 acts mechanically as part of the Polycomb group (PcG) proteins. It weighs approximately 60 kDa and it is found in nuclear matrix where it participates in chromatin remodeling. CBX4 shows significant expression in tissues like liver brain and lung indicating its wide importance to cellular function. By binding to methylated histones especially H3K27me3 CBX4 influences gene silencing and epigenetic regulation important for maintaining cellular identity.
CBX4 operates as a component of the Polycomb Repressive Complex 1 (PRC1) which plays role in controlling gene expression patterns during development. In this complex CBX4 interacts with other core components such as BMI1 and RING1 to facilitate monoubiquitination of histone H2A leading to transcriptional repression. This ability of CBX4 to modulate epigenetic states highlights its importance in critical developmental processes and differentiation.
CBX4 significantly influences gene regulatory networks involved in cell cycle and apoptosis pathways. In the cell cycle regulation CBX4 interacts with important proteins such as Cyclin D1 impacting cell proliferation. In apoptosis CBX4 has roles linked to the p53 pathway by modulating transcriptional activity which influences cellular fate. These pathways underline the significance of CBX4 in maintaining balance between cell growth and death.
The deregulation of CBX4 expression has been associated with cancers like hepatocellular carcinoma and breast cancer. CBX4 potentially interacts with proteins such as p21 and p16 whose pathways play significant roles in cancer development. This underlines its role as a potential biomarker or therapeutic target in cancer treatment strategies. Understanding the function and regulation of CBX4 could offer insights in devising novel treatments for these malignancies.
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False colour image of Western blot: Anti-CBX4 antibody [EPR23053-7] - ChIP Grade 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-CBX4 antibody [EPR23053-7] - ChIP Grade ab242149 was shown to bind specifically to CBX4. A band was observed at 75 kDa in wild-type HEK-293 cell lysates with no signal observed at this size in Cbx4 knockout cell line ab261723 (knockout cell lysate ab261666). The band observed in the knockout lysate lane below 75 kDa is likely to represent a truncated form of CBX4. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and Cbx4 knockout HEK-293 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°C. 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-CBX4 antibody [EPR23053-7] - ChIP Grade (Anti-CBX4 antibody [EPR23053-7] - ChIP Grade ab242149) at 1/1000 dilution
Lane 1: Wild-type HEK-293 cell lysate at 20 µg
Lane 2: Cbx4 CRISPR-Cas9 edited HEK-293 cell lysate at 20 µg
Lane 2: Western blot - Human CBX4 knockout HEK-293 cell line (ab261723)
Lane 3: RAW 264.7 cell lysate at 20 µg
Lane 4: HUVEC cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 61 kDa
Observed band size: 75 kDa
Knockout achieved by CRISPR/Cas9; X = 7 bp deletion, 1 bp insertion, 10 bp deletion
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