TREX1 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 insertion in exon 1 and 4 bp insertion in exon 1.
3' 5' exonuclease TREX1, 3' repair exonuclease 1, AGS1, AGS5, CRV, DKFZp434J0310, DNase III, DRN3, Deoxyribonuclease III, dnaQ/mutD (E. coli) like, HERNS, Three prime repair exonuclease 1
TREX1 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 insertion in exon 1 and 4 bp insertion in exon 1.
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.
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.
TREX1 also known as Three Prime Repair Exonuclease 1 acts as a DNA exonuclease which degrades single-stranded DNA from the 3' end. This protein exhibits a mass of 33 kDa. TREX1's expression is largely found in the cytoplasm of many cell types including those in the immune system and various tissues like the liver spleen and heart. Through its enzymatic activity TREX1 helps maintain genomic stability by processing DNA intermediates that arise from incomplete DNA replication or repair processes.
TREX1 removes excessive DNA that might trigger immune responses. It is not a part of a large protein complex but works closely with substrates involved in DNA repair and replication. By degrading abnormal DNA TREX1 prevents the initiation of inappropriate immune responses that could result in autoimmunity. Its functionality is essential in preventing the cell from converting these DNA fragments into ligands for DNA sensors which could activate immune signaling pathways.
TREX1 plays a significant role in the DNA damage response and innate immune pathways. In the DNA damage response pathway TREX1 functions alongside proteins like ATR and ATM which address DNA replication stress and repair. Within the innate immune pathway it interacts with cGAS (cyclic GMP-AMP synthase) which can become activated in response to cytosolic DNA leading to the production of type I interferons - powerful immune modulators.
Mutations in TREX1 have been linked to autoimmune diseases such as Aicardi-Goutières syndrome and systemic lupus erythematosus. In these conditions the altered or deficient TREX1 leads to the accumulation of DNA in the cytoplasm falsely triggering an antiviral immune response. This relationship with cGAS is critical as the cGAS-STING pathway becomes activated due to the presence of unprocessed DNA resulting in chronic inflammation and contributing to the autoimmune pathology observed in these disorders.
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Anti-TREX1 antibody [EPR14985] ab185228 Anti-TREX1 antibody [EPR14985] was shown to specifically react with TREX1 in wild-type A549 cells. Loss of signal was observed when knockout cell line ab266927 (knockout cell lysate Human TREX1 knockout A549 cell lysate ab257764) was used. Wild-type and TREX1 knockout samples were subjected to SDS-PAGE. Anti-TREX1 antibody [EPR14985] ab185228 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) 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-TREX1 antibody [EPR14985] (Anti-TREX1 antibody [EPR14985] ab185228) at 1/1000 dilution
Lane 1: Wild-type A549 cell lysate at 20 µg
Lane 2: TREX1 knockout A549 cell lysate at 20 µg
Lane 2: Western blot - Human TREX1 knockout A549 cell line (ab266927)
Lane 3: Raji cell lysate at 20 µg
Lane 4: Daudi 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: 14 kDa, 39 kDa
Observed band size: 14 kDa, 34 kDa
Representative images of TREX1 knockout A549 cells, low and high confluency examples (top left and right respectively) and wild-type A549 cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.
Allele-2: 4 bp insertion in exon 1.
Allele-1: 1 bp insertion in exon1
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