GSK3A KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and Insertion of the selection cassette in exon 1.
HEK-293T
Human
Kidney
Liquid
Sanger Sequencing, Western blot
Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and Insertion of the selection cassette in exon 1
DKFZp686D0638, GSK-3 alpha, GSK3A_HUMAN, Glycogen synthase kinase-3 alpha, Serine/threonine protein kinase GSK3A
GSK3A KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and Insertion of the selection cassette in exon 1.
HEK-293T
Human
Kidney
Liquid
Sanger Sequencing, Western blot
Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 1 and Insertion of the selection cassette in exon 1
GSK3A
Knockout
CRISPR technology
Sanger Sequencing, Western blot
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
EU: 2 US: 2
Adherent
Female
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.
DMEM (High Glucose) + 10% FBS
Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Dry Ice
-196°C
-196°C
Recommended control: Human wild-type HEK293T cell line (Human wild-type HEK-293T 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.
This supplementary information is collated from multiple sources and compiled automatically.
Glycogen synthase kinase 3 alpha (GSK3 alpha) is a serine/threonine protein kinase also known as GSK3A with a molecular mass of approximately 51 kDa. It plays an important role in the regulation of various cellular processes. GSK3 alpha is expressed in many tissues but has high levels in the brain heart and skeletal muscle. It participates in the phosphorylation of numerous substrates impacting their activity and stability.
GSK3 alpha influences several cellular functions including cell division differentiation and apoptosis. It functions independently or as part of a protein complex often forming dimeric complexes with GSK3 beta. GSK3 alpha alters the activity of numerous transcription factors and regulates insulin and Wnt signaling pathways affecting glucose metabolism and cellular growth.
GSK3 alpha is involved in the Wnt and insulin signaling pathways. In the Wnt pathway GSK3 alpha phosphorylates beta-catenin marking it for degradation and controlling gene expression. In the insulin pathway it modulates glycogen synthesis and affects glucose homeostasis. GSK3 alpha interacts closely with proteins like APC and glycogen synthase making it integral to these pathways.
GSK3 alpha links to neurodegenerative disorders and diabetes. Its dysfunction contributes to Alzheimer’s disease where abnormal phosphorylation of tau protein occurs. In the context of diabetes GSK3 alpha influences glucose regulation impacting insulin sensitivity and action. The interplay between GSK3 alpha and tau in Alzheimer's as well as insulin signaling proteins illustrates its important role in these disease pathways.
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Lanes 1-4: Merged signal (red and green). Green - Anti-GSK3 alpha antibody [EP793Y] ab40870 observed at 51 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-GSK3 alpha antibody [EP793Y] ab40870 Anti-GSK3 alpha antibody [EP793Y] was shown to specifically react with GSK3 alpha in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266742 (knockout cell lysate Human GSK3A (GSK3 alpha) knockout HEK-293T cell lysate ab257979) was used. Wild-type and GSK3 alpha knockout samples were subjected to SDS-PAGE. Anti-GSK3 alpha antibody [EP793Y] ab40870 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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-GSK3 alpha antibody [EP793Y] (Anti-GSK3 alpha antibody [EP793Y] ab40870) at 1/1000 dilution
Lane 1: Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2: GSK3A knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4: HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg
All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 51 kDa
Observed band size: 51 kDa
This data was developed using Anti-GSK3 alpha antibody [EP793Y] ab40870, the same antibody clone in a different buffer formulation.
Lanes 1-4: Merged signal (red and green). Green - Anti-GSK3 alpha antibody [EP793Y] ab40870 observed at 51 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-GSK3 alpha antibody [EP793Y] ab40870 Anti-GSK3 alpha antibody [EP793Y] was shown to specifically react with GSK3 alpha in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266742 (knockout cell lysate Human GSK3A (GSK3 alpha) knockout HEK-293T cell lysate ab257979) was used. Wild-type and GSK3 alpha knockout samples were subjected to SDS-PAGE. Anti-GSK3 alpha antibody [EP793Y] ab40870 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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.
Lanes 1-4: Merged signal (red and green). Green - Anti-GSK3 alpha antibody ab78664 observed at 51 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-GSK3 alpha antibody ab78664 Anti-GSK3 alpha antibody was shown to specifically react with GSK3 alpha in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266742 (knockout cell lysate Human GSK3A (GSK3 alpha) knockout HEK-293T cell lysate ab257979) was used. Wild-type and GSK3 alpha knockout samples were subjected to SDS-PAGE. Anti-GSK3 alpha antibody ab78664 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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-GSK3 alpha antibody (Anti-GSK3 alpha antibody ab78664) at 1/1000 dilution
Lane 1: Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2: GSK3A knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4: HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg
All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 51 kDa
Observed band size: 51 kDa
Lanes 1-4: Merged signal (red and green). Green - Anti-GSK3 alpha antibody ab78664 observed at 51 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-GSK3 alpha antibody ab78664 Anti-GSK3 alpha antibody was shown to specifically react with GSK3 alpha in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266742 (knockout cell lysate Human GSK3A (GSK3 alpha) knockout HEK-293T cell lysate ab257979) was used. Wild-type and GSK3 alpha knockout samples were subjected to SDS-PAGE. Anti-GSK3 alpha antibody ab78664 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (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.
Representative images GSK3A 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 M5000 microscope.
Allele-1: 1 bp deletion in exon1
Allele-2: Insertion of the selection cassette in exon 1.
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