Human H2AFY (mH2A1) knockout HEK-293T cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human H2AFY (mH2A1) knockout HEK-293T cell line (AB266241)
Lanes 1- 2 : Merged signal (red and green). Green - ab183041 observed at 40 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.
ab183041 was shown to react with mH2A1 in wild-type HEK-293T cells in western blot. The band observed in CRISPR/Cas9 edited cell line ab266241 (CRISPR/Cas9 edited cell lysate ab257463) lane below 40kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HEK-293T and H2AFY CRISPR/Cas9 edited HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab183041 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4° at a 1 in 1000 Dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-mH2A1 antibody [EPR9359(2)] (<a href='/en-us/products/primary-antibodies/mh2a1-antibody-epr93592-ab183041'>ab183041</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
H2AFY CRISPR/Cas9 edited HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human H2AFY (mH2A1) knockout HEK-293T cell line (ab266241)
Predicted band size: 40 kDa
Observed band size: 40 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human H2AFY (mH2A1) knockout HEK-293T cell line (AB266241)
Homozygous : 13 bp deletion in exon 2
- Cell Culture
Unknown
Cell Culture - Human H2AFY (mH2A1) knockout HEK-293T cell line (AB266241)
Representative images of H2AFY knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS XL Core microscope.
Complementary Products
![Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-mH2A1 antibody [EPR9359(2)] (AB183041)](https://content.abcam.com/products/images/mh2a1-antibody-epr93592-ab183041--immunohistochemistry-formalin-pfa-fixed-paraffin-embedded-sections-img131563.jpg)
Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant 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
Zygosity
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
MH2A1 plays significant roles in chromatin remodeling and gene expression regulation. It is part of the nucleosome replacing canonical histone H2A and influencing the accessibility of chromatin to transcriptional machinery. Through this function mH2A1 can repress or activate gene expression depending on cellular context and interacting partners. Its incorporation into chromatin is associated with the silencing of certain genomic regions such as inactive X chromosome in females with potential implications for epigenetic modifications and cellular differentiation.
Pathways
MH2A1 is notably involved in pathways regulating chromatin organization and cell differentiation. It plays a role in the Polycomb Repressive Complex 2 (PRC2) pathway affecting histone methylation and gene silencing. Furthermore mH2A1 interacts with proteins like EZH2 in PRC2 which carry out methylation at histone H3 on lysine 27. These interactions highlight mH2A1's involvement in maintaining repressive chromatin states and controlling gene expression patterns important for normal cell function.
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
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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