Human NNMT knockout HeLa cell line
- Advanced Validation
- What is this?
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NNMT KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 16 bp deletion in exon 1 and 1 bp insertion in exon 1.
View Alternative Names
EC 2.1.1.1, NNMT_HUMAN, Nicotinamide N-methyltransferase
- WB
Lab
Western blot - Human NNMT knockout HeLa cell line (AB265700)
Lanes 1-3 : Merged signal (red and green). Green - ab119758 observed at 30 kDa. Red - loading control ab181602 observed at 36 kDa.
ab119758 Anti-NNMT antibody [OTI3D8] was shown to specifically react with NNMT in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265700 (knockout cell lysate ab258537) was used. Wild-type and NNMT knockout samples were subjected to SDS-PAGE. ab119758 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated at room temperature for 2.5 hours at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-NNMT antibody [OTI3D8] (<a href='/en-us/products/primary-antibodies/nnmt-antibody-oti3d8-ab119758'>ab119758</a>) at 1/500 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
NNMT knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human NNMT knockout HeLa cell line (ab265700)
Lane 3:
A549 cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-680rd-preadsorbed-ab216777'>ab216777</a>) at 1/10000 dilution
Predicted band size: 30 kDa
Observed band size: 30 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human NNMT knockout HeLa cell line (AB265700)
Allele-2 : 1 bp insertion in exon 1.
- Sanger seq
Unknown
Sanger Sequencing - Human NNMT knockout HeLa cell line (AB265700)
Allele-1 : 16 bp deletion in exon 1.
Reactivity data
Product details
Recommended control: Human wild-type HeLa cell line (ab255928). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
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
NNMT impacts cell energy balance and gene expression regulation by influencing the levels of nicotinamide and methionine metabolites. This enzyme does not typically form part of a complex acting more so as an individual regulator of methylation reactions affecting cellular metabolism. Researchers find interest in NNMT for its role in modulating cellular growth and proliferation forming a link to both normal cellular functions and pathological conditions like cancer.
Pathways
NNMT has a significance in NAD+ metabolism and methionine salvage pathways. These pathways are essential in maintaining redox homeostasis and proper methylation status in cells. NNMT through its enzymatic activity interacts with SAM (S-adenosyl methionine) closely relating its function to methylation processes that involve other proteins such as SIRTs (Sirtuins) and NAMPT (Nicotinamide phosphoribosyltransferase) within these metabolic pathways.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Complementary Products
![Western blot - Anti-NNMT antibody [OTI3D8] (AB119758)](https://content.abcam.com/products/images/nnmt-antibody-oti3d8-ab119758--western-blot-img5921.jpg)
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