Human PUS1 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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Human PUS1 knockout HEK-293T cell line available to order. Recommended control: Human wild-type HEK293T cell line (ab255449).
View Alternative Names
A730013B20Rik, DOBI, MGC112655, MGC11268, MLASA, MLASA1, Pseudouridine synthase 1, Pseudouridylate synthase 1, TRUA_HUMAN, mPus1p, tRNA pseudouridine synthase A, mitochondrial, tRNA pseudouridine(38-40) synthase, tRNA pseudouridylate synthase I, tRNA-uridine isomerase I
- WB
Lab
Western blot - Human PUS1 knockout HEK-293T cell line (AB266091)
Lanes 1- 4 : Merged signal (red and green). Green - ab203010 observed at 45 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.
ab203010 was shown to react with PUS1 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266091 (knockout cell lysate ab258158) was used. Wild-type HEK-293T and PUS1 knockout HEK-293T cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab203010 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4° at a 1 in 1000 dilution and a 1 in 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 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-PUS1 antibody [EPR20181] (<a href='/en-us/products/primary-antibodies/pus1-antibody-epr20181-ab203010'>ab203010</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK-293T cell lysate at 20 µg
Lane 2:
PUS1 knockout HEK-293T cell lysate at 20 µg
Lane 2:
Western blot - Human PUS1 knockout HEK-293T cell line (ab266091)
Lane 3:
HeLa cell lysate at 20 µg
Lane 4:
Daudi cell lysate at 20 µg
Predicted band size: 47 kDa
Observed band size: 45 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human PUS1 knockout HEK-293T cell line (AB266091)
Allele-2 : 41 bp deletion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human PUS1 knockout HEK-293T cell line (AB266091)
Allele-1 : 47 bp deletion in exon2
Reactivity data
Product details
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 modification of RNA by PUS1 improves the stability and folding of the RNA structure aiding its functionality. The enzyme is not part of a larger complex but acts independently to achieve its role in RNA modification. Pseudouridine created by PUS1 permits enhanced base stacking and hydrogen bonding which influences the translation process and the overall efficiency of protein synthesis. This modification is important for the proper functioning of tRNA and rRNA ensuring fidelity during protein translation.
Pathways
PUS1 plays an essential role in the RNA processing and modification pathways. Its activity integrates into pathways such as ribosome biogenesis and RNA quality control. Particularly it interacts with other RNA-binding proteins and enzymes like TRUB1 another pseudouridine synthase which also contributes to the maturation of structured RNAs. These pathways are vital for maintaining cellular homeostasis and responding to various physiological demands.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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