DNMT3A Knockout HeLa Cell Lysate

Product Discontinued

DNMT3A Knockout HeLa Cell Lysate Summary

• Preparation Method: Knockout achieved by using CRISPR/Cas9,-1 bp deletion in exon and 1 bp insertion in exon
• Gene: DNMT3A

Applications/Dilutions

• Dilutions:
  • Western Blot
• Application Notes: You will receive 1 vial (100ug) of knockout cell lysate and 1 vial (100ug) of Parental cell lysate. Lysate can be diluted with 1X SDS sample buffer and will be stable at -20 degrees C for 12 months. Minimize freeze-thaw cycles.

Packaging, Storage & Formulations

• Storage: Store at -20C short term. Aliquot and store at -80C long term. Avoid freeze-thaw cycles.
• Buffer: 0.1 mg cell homogenate lyophilized in RIPA buffer made with double-knockout cell lines.
• Concentration: LYOPH
• Reconstitution Instructions: To use as WB negative control, spin down briefly and resuspend in 100 uL 1xSDS sample buffer (2% SDS, 60 mM Tris-HCl pH 6.8, 10% Glycerol, 0.02% Bromophenol blue, 60 mM beta-mercaptoethanol). Boil the lysate for 3 - 5 minutes before loading it onto gel.

Lysate Details for Array

• Type: Knockout HeLa Cell

Notes

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Validation of antibody specificity is critical and verification of antibody performance against knockout samples is one way to guarantee that an antibody recognizes a specific target. Novus' KO cell lysate can be used as a negative control for western blots and to confirm the specificity of antibodies.

Alternate Names for DNMT3A Knockout HeLa Cell Lysate

• DNA (cytosine-5-)-methyltransferase 3 alpha
• DNA (cytosine-5)-methyltransferase 3A
• DNA cytosine methyltransferase 3A2
• DNA Methyltransferase 3 Alpha
• DNA methyltransferase HsaIIIA
• DNA MTase HsaIIIA
• DNMT3A
• DNMT3A2
• EC 2.1.1.37
• M.HsaIIIA
• TBRS

Background

Methylation of cytosine residues is essential for mammalian development by regulating gene expression, gene silencing and genomic imprinting (1). This process is controlled by the enzyme DNA (cytosine-5)-methyltransferase 3A (DNMt3a), which is encoded by the DNMT3A gene and has a predicted molecular weight of 102 kDa. DNA methyltransferases including DNMt3a are involved in de novo cytosine methylation and maintenance of embryonic and somatic cells through the transfer of methyl groups to specific CpG structures in DNA (2). Somatic mutations in DNMT3A are responsible for cytogenetically normal acute myeloid leukemia (CN-AML). Hypermethylation of CpG islands in tumor suppressor genes or hypomethylation of bulk genomic DNA are linked with cancer (3,4). Mutations in the DNMT3A gene have also been found to cause DNMT3A overgrowth syndrome and myelodysplastic syndromes.

References

1. Thakur, A., Mackin, S. J., Irwin, R. E., O'Neill, K. M., Pollin, G., & Walsh, C. (2016). Widespread recovery of methylation at gametic imprints in hypomethylated mouse stem cells following rescue with DNMT3A2. Epigenetics Chromatin, 9, 53. doi:10.1186/s13072-016-0104-2

2. Ravichandran, M., Lei, R., Tang, Q., Zhao, Y., Lee, J., Ma, L., . . . Dawlaty, M. M. (2019). Rinf Regulates Pluripotency Network Genes and Tet Enzymes in Embryonic Stem Cells. Cell Rep, 28(8), 1993-2003.e1995. doi:10.1016/j.celrep.2019.07.080

3. Pang, Y., Liu, J., Li, X., Xiao, G., Wang, H., Yang, G., . . . Ren, H. (2018). MYC and DNMT3A-mediated DNA methylation represses microRNA-200b in triple negative breast cancer. J Cell Mol Med, 22(12), 6262-6274. doi:10.1111/jcmm.13916

4. Xia, L., Huang, W., Bellani, M., Seidman, M. M., Wu, K., Fan, D., . . . Baylin, S. B. (2017). CHD4 Has Oncogenic Functions in Initiating and Maintaining Epigenetic Suppression of Multiple Tumor Suppressor Genes. Cancer Cell, 31(5), 653-668.e657. doi:10.1016/j.ccell.2017.04.005

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Lysates are guaranteed for 6 months from date of receipt.

Bioinformatics

• Gene Symbol: DNMT3A