Human SPHK2 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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SPHK2 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 1.
View Alternative Names
SK 2, SPHK2_HUMAN, SPK 2, Sphingosine kinase 2, Sphingosine kinase type 2
- WB
Lab
Western blot - Human SPHK2 knockout HEK-293T cell line (AB267293)
Lanes 1 - 4 : Merged signal (red and green). Green - Anti-SPHK2 antibody observed at 68 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37 kDa.
Anti-SPHK2 antibody was shown to react with SPHK2 in wild-type HEK-293T cells in Western blot with loss of signal observed in SPHK2 knockout cell line ab267293 (SPHK2 knockout cell lysate ab258699). Wild-type HEK-293T and SPHK2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with anti-SPHK2 antibody and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4 °C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
All lanes:
Anti-SPHK2 antibody at 1/1000 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
SPHK2 knockout HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human SPHK2 knockout HEK-293T cell line (ab267293)
Lane 3:
HeLa cell lysate at 20 µg
Lane 4:
Raji cell lysate at 20 µg
false
- Sanger seq
Unknown
Sanger Sequencing - Human SPHK2 knockout HEK-293T cell line (AB267293)
Homozygous : 1 bp insertion in exon1
Reactivity data
Product details
Recommended control: Human wild-type HEK293T cell line (ab255449). 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
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
SPHK2 influences cell growth survival and differentiation by generating sphingosine-1-phosphate a signaling lipid. It functions independently or in association with other proteins but it is not always a part of a larger complex. The generated S1P acts as a signaling molecule both inside and outside of cells affecting processes like apoptosis and inflammation.
Pathways
SPHK2 is a significant player in the sphingolipid signaling pathway and impacts the MAPK/ERK pathway. These pathways collectively regulate cellular proliferation and apoptosis. SPHK2 works in conjunction with SPHK1 a closely related sphingosine kinase in these signaling routes. These kinases together help to balance the levels of sphingosine and S1P which is essential for maintaining cellular homeostasis.
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
Complementary Products
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