Human TRMT1 (TRM1) knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
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 - ab283652 observed at 75 kDa. Red - loading control ab8245 observed at 36 kDa. 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 ab258252) was used. Wild-type and TRMT1 (TRM1) knockout samples were subjected to SDS-PAGE. ab283652 and 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 (ab216773) and 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] (<a href='/en-us/products/primary-antibodies/trm1-antibody-epr25054-72-ab283652'>ab283652</a>) 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
Secondary
All lanes:
Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (<a href='/en-us/products/secondary-antibodies/goat-mouse-igg-h-l-irdye-680rd-preadsorbed-ab216776'>ab216776</a>), 1/10000 dilution
Predicted band size: 72 kDa
Observed band size: 75 kDa
false
- WB
Lab
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] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, 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 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
All lanes:
Western blot - Anti-TRM1 antibody - C-terminal (<a href='/en-us/products/primary-antibodies/trm1-antibody-c-terminal-ab185997'>ab185997</a>) 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
Predicted band size: 72 kDa
Observed band size: 75 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human TRMT1 (TRM1) knockout HEK-293T cell line (AB266705)
Allele-2 : Insertion of the selection cassette in exon 6.
- Cell Culture
Unknown
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 seq
Unknown
Sanger Sequencing - Human TRMT1 (TRM1) knockout HEK-293T cell line (AB266705)
Allele-1 : Insertion of the selection cassette in exon 6
Complementary Products
Primary Antibodies
AB283652
Anti-TRM1 antibody [EPR25054-72]
20ul selling size|RabMAb|Recombinant|KO Validated|Trial Size
![Western blot - Anti-TRM1 antibody [EPR25054-72] (AB283652)](https://content.abcam.com/products/images/trm1-antibody-epr25054-72-ab283652--western-blot-img170628.jpg)
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Primary Antibodies
AB281926
Anti-Eph receptor A3 antibody [EPR24479-136]
20ul selling size|RabMAb|Recombinant
![Western blot - Anti-Eph receptor A3 antibody [EPR24479-136] (AB281926)](https://content.abcam.com/products/images/eph-receptor-a3-antibody-epr24479-136-ab281926--western-blot-img176770.jpg)
- 0
- 0
Reactivity data
Product details
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
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
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.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
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