TRMT1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Insertion of the selection cassette in exon 6.
Images
Key facts
- Cell type
- HEK-293T
- Species or organism
- Human
- Tissue
- Kidney
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Insertion of the selection cassette in exon 6
Alternative names
2)G26)dimethyltransferase, 2-dimethylguanosine-26 methyltransferase, FLJ20244, N(2) N(2) dimethylguanosine tRNA methyltransferase, N(2)-N(2)) methyltransferase, TRM1 tRNA methyltransferase 1 homolog, TRM1_HUMAN, TRMT 1, tRNA (guanine(26)-N(2))-dimethyltransferase, tRNA 2, tRNA 2 2 dimethylguanosine 26 methyltransferase, tRNA(guanine 26 N(2) N(2)) methyltransferase, tRNA(guanine-26, tRNA(m(2, tRNA(m(2 2)G26)dimethyltransferase
Recommended products
TRMT1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Insertion of the selection cassette in exon 6.
Key facts
- Cell type
- HEK-293T
- Species or organism
- Human
- Tissue
- Kidney
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Insertion of the selection cassette in exon 6
- Concentration
Properties
- Gene name
- TRMT1
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing, Western blot
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 2 US: 2
- Adherent/suspension
- Adherent
- Gender
- Female
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.
Storage
- Shipped at conditions
- Dry Ice
- Appropriate short-term storage conditions
- -196°C
- Appropriate long-term storage conditions
- -196°C
Notes
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.
Supplementary info
- This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
TRM1 also known as transfer RNA methyltransferase 1 is an enzyme responsible for methylating the N2 position of guanine in tRNA molecules. It has a molecular mass of approximately 49 kDa. TRM1 mostly resides in the mitochondria but can also be found in the cytoplasm. Its presence ensures proper methylation of tRNA influencing efficient and accurate protein translation.
- Biological function summary
The methylation activity of TRM1 plays a role in maintaining the stability and function of tRNA molecules. As part of the cellular machinery TRM1 does not function alone; it often works together with other tRNA-modifying enzymes to ensure the fidelity of protein synthesis. This modification protects tRNA's structure facilitates proper folding and enhances interaction with ribosomal components.
- Pathways
TRM1 functions significantly impact protein synthesis pathways. The enzyme is instrumental in the translation process by ensuring that tRNA molecules are correctly modified for protein assembly. TRM1 operates alongside other proteins such as TRM10 and TRMT5 within these modification pathways influencing the efficiency and accuracy of translation. It supports processes like tRNA maturation and ribosomal activity.
- Associated diseases and disorders
Alterations in TRM1 activity can relate to mitochondrial pathologies. Mutations or deficiencies in TRM1 can disrupt mitochondrial function leading to disorders such as mitochondrial myopathy. Research also connects TRM1 aberrations to neurodegenerative diseases where it may interact with proteins like PINK1 highlighting its role in neuroprotective responses. The efficiency of TRM1 in performing its tasks is vital as any dysfunction can result in profound cellular consequences affecting human health.
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5 product images
Western blot - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
False colour image of Western blot: Anti-TRM1 antibody - C-terminal staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-TRM1 antibody - C-terminal ab185997 was shown to bind specifically to TRM1. A band was observed at 75 kDa in wild-type HEK-293T cell lysates with no signal observed at this size in Trmt1 knockout cell line ab266705 (knockout cell lysate Human TRMT1 (TRM1) knockout HEK-293T cell lysate ab258252). To generate this image, wild-type and Trmt1 knockout HEK-293T cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. 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 (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-TRM1 antibody - C-terminal (Anti-TRM1 antibody - C-terminal ab185997) at 1/1000 dilution
Lane 1: Wild-type HEK-293T cell lysate at 20 µg
Lane 2: TRMT1 knockout HEK-293T cell lysate at 20 µg
Lane 2: Western blot - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Lane 3: HeLa cell lysate at 20 µg
Lane 4: HepG2 cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 72 kDa
Observed band size: 75 kDa
Western blot - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Blocking and diluting buffer and concentration: Intercept®(TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS.
Lanes 1-3:Merged signal (red and green). Green - Anti-TRM1 antibody [EPR25054-72] ab283652 observed at 75 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa. Anti-TRM1 antibody [EPR25054-72] ab283652 Anti-TRMT1 (TRM1) antibody [EPR25054-72] was shown to specifically react with TRMT1 (TRM1) in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266705 (knockout cell lysate Human TRMT1 (TRM1) knockout HEK-293T cell lysate ab258252) was used. Wild-type and TRMT1 (TRM1) knockout samples were subjected to SDS-PAGE. Anti-TRM1 antibody [EPR25054-72] ab283652 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 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 1/10000 dilution for 1 hour at room temperature before imaging.
The bands nearby 100 kDa could be non-specific bands.
All lanes: Western blot - Anti-TRM1 antibody [EPR25054-72] (Anti-TRM1 antibody [EPR25054-72] ab283652) at 1/1000 dilution
Lane 1: Wild-type HEK293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg
Lane 2: TRMT1 (TRM1) knockout HEK293T (ab266705) whole cell lysate at 20 µg
Lane 2: Western blot - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Lane 3: HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate at 20 µg
SecondaryAll lanes: Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776), 1/10000 dilution
Predicted band size: 72 kDa
Observed band size: 75 kDa
Cell Culture - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Representative images of TRMT1 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 XL Core microscope.
Sanger Sequencing - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Allele-1: Insertion of the selection cassette in exon 6
Sanger Sequencing - Human TRMT1 (TRM1) knockout HEK-293T cell line (ab266705)
Allele-2: Insertion of the selection cassette in exon 6.
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