Human DEPDC5 knockout A549 cell line
- Advanced Validation
- What is this?
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DEPDC5 KO cell line available to order. KO validated by. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 12.
View Alternative Names
DEP domain containing 5, DEP domain-containing protein 5, DEP.5, DEPD5_HUMAN, FFEVF, KIAA0645
- WB
Lab
Western blot - Human DEPDC5 knockout A549 cell line (AB266906)
False colour image of Western blot : Anti-DEPDC5 antibody [EPR20497-23] staining at 1/1000 dilution shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution shown in red. In Western blot ab213181 was shown to bind specifically to DEPDC5. A band was observed at 181 kDa in wild-type A549 cell lysates with no signal observed at this size in DEPDC5 knockout cell line ab266906 (knockout cell lysate ab258394). To generate this image wild-type and DEPDC5 knockout A549 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged.Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
All lanes:
Western blot - Anti-DEPDC5 antibody [EPR20497-23] (<a href='/en-us/products/primary-antibodies/depdc5-antibody-epr20497-23-ab213181'>ab213181</a>) at 1/1000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
DEPDC5 knockout A549 cell lysate at 20 µg
Lane 2:
Western blot - Human DEPDC5 knockout A549 cell line (ab266906)
Lane 3:
Human brain tissue lysate at 20 µg
Lane 4:
Mouse brain tissue lysate at 20 µg
Predicted band size: 178 kDa
Observed band size: 181 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human DEPDC5 knockout A549 cell line (AB266906)
Homozygous : 1 bp insertion in exon12
Reactivity data
Product details
Recommended control: Human wild-type A549 cell line (ab255450). 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.
- Do not allow the cell density to exceed 7x104 cells/cm2.
Culture medium
F-12K + 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
DEPDC5 acts as a negative regulator of the mTORC1 signaling pathway by being a part of the GATOR1 complex. This complex is involved in nutrient sensing and regulation of cell growth and metabolism. DEPDC5's role in mTORC1 regulation links it to the control of protein synthesis and autophagy essential processes in maintaining cellular homeostasis. By interacting with other components of the GATOR1 complex DEPDC5 contributes not only to inhibiting mTORC1 activity but also to the broader metabolic balance.
Pathways
DEPDC5 is integral to the mTOR signaling pathway and the amino acid sensing pathways. The protein acts in coordination with other GATOR1 complex components such as NPRL2 and NPRL3 which together monitor nutrient availability and adapt cellular functions accordingly. These pathways highlight DEPDC5's role in connecting nutrient signals to cellular growth responses showing its influence on critical regulatory networks affecting cell growth and proliferation.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
Target data
Complementary Products
![Western blot - Anti-DEPDC5 antibody [EPR20497-23] (AB213181)](https://content.abcam.com/products/images/depdc5-antibody-epr20497-23-ab213181--western-blot-img29227.jpg)
Primary Antibodies
AB105459
BOND RX™ Validated|KO Validated|Trial Size
![Western blot - Anti-WIPI2 antibody [2A2] (AB105459)](https://content.abcam.com/products/images/wipi2-antibody-2a2-ab105459--western-blot-img1726.jpg)
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Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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