RHOT2 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 2 and 2 bp deletion in exon 2.
Images
Key facts
- Cell type
- HeLa
- Species or organism
- Human
- Tissue
- Cervix
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 2 and 2 bp deletion in exon 2
Alternative names
ARHT2, C16orf39, MIRO2_HUMAN, Mitochondrial Rho GTPase 2, RASL, RHOT2, Ras homolog gene family member T2, hMiro-2
Recommended products
RHOT2 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 2 and 2 bp deletion in exon 2.
Key facts
- Cell type
- HeLa
- Species or organism
- Human
- Tissue
- Cervix
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Knockout achieved by using CRISPR/Cas9, 1 bp deletion in exon 2 and 2 bp deletion in exon 2
- Disease
- Adenocarcinoma
- Concentration
Properties
- Gene name
- RHOT2
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing
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 and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant 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
MIRO2 also known as Rhot2 or Mitochondrial Rho GTPase 2 is a mitochondrial Rho GTPase with a molecular weight of approximately 69 kDa. This protein belongs to the Rho family of GTPases and exists mainly on the mitochondrial outer membrane. It acts as a critical regulator of mitochondrial trafficking and dynamics. MIRO2 plays a pivotal role in the regulation of mitochondrial transport along the cytoskeleton by interacting with adaptor proteins. It features two EF-hand motifs and two GTPase domains that are essential for its function.
- Biological function summary
MIRO2 contributes to the maintenance of mitochondrial network and quality within cells. It serves as a component of the calcium-sensing mitochondrial transport machinery and interacts with the transport protein complex including adaptor proteins like Milton and the motor protein kinesin. Through these interactions MIRO2 helps in coupling mitochondrial dynamics to cellular events by facilitating calcium-dependent docking and release of mitochondria. This function indicates the importance of MIRO2 in energy-demanding cellular processes.
- Pathways
MIRO2 integrates into the mitochondrial transport and positioning pathways. It associates with the PINK1/Parkin pathway which is essential for mitochondrial quality control and turnover. The pathway leads to the balance of mitochondrial fission and fusion assisting in the removal of damaged mitochondria. Moreover MIRO2 collaborates with proteins like Mitofusins in these pathways to ensure efficient mitochondrial dynamics and bioenergetics.
- Associated diseases and disorders
MIRO2 has links to neurodegenerative diseases like Parkinson's disease. Dysfunction in the PINK1/Parkin pathway where MIRO2 participates leads to impaired mitophagy and neuronal cell death. Additionally imbalances in MIRO2 activity are implicated in the progression of certain types of cancer. In these contexts MIRO2 interacts with related proteins such as PINK1 and Mitofusins which play roles in disease mechanisms that involve mitochondrial dysfunction or aberrant cellular metabolism.
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4 product images
Sanger Sequencing - Human RHOT2 (MIRO2) knockout HeLa cell line (ab265801)
Allele-2: 1 bp deletion in exon 2.
Sanger Sequencing - Human RHOT2 (MIRO2) knockout HeLa cell line (ab265801)
Allele-1: 2 bp deletion in exon 2.
Immunocytochemistry/ Immunofluorescence - Human RHOT2 (MIRO2) knockout HeLa cell line (ab265801)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized RHOT2 KO HeLa (RHOT2 knockout human cervical adenocarcinoma epithelial cell), ab265801 cells labelling MIRO2 with Anti-MIRO2 antibody [RM2077] ab322962 at 1/500 (0.906 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).
Confocal image showing mitochondrial staining in wildtype HeLa cells (shown in green), showing no staining in RHOT2 knockout HeLa cells. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
anti-COX IV mouse monoclonal antibody - Mitochondrial Marker was used to counterstain tubulin at 1/200 2.5 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).
-ve control 1: Anti-MIRO2 antibody [RM2077] ab322962 at 1/500 dilution, followed byab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution.
-ve control 2: anti-COX IV mouse monoclonal - Mitochondrial Marker antibody at 1/200 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) antibody at 1/1000 dilution.
Immunocytochemistry/ Immunofluorescence - Human RHOT2 (MIRO2) knockout HeLa cell line (ab265801)
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized RHOT2 KO HeLa (RHOT2 knockout human cervical adenocarcinoma epithelial cell), ab265801 cells labelling MIRO2 with Anti-MIRO2 antibody [EPR29118-85] ab320739 at 1/50 (9.12 ug/ml) dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).
Confocal image showing mitochondrial staining in wildtype HeLa cells (shown in green), showing no staining in RHOT2 knockout HeLa cells. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
anti-COX IV mouse monoclonal antibody - Mitochondrial Marker was used to counterstain tubulin at 1/200 (2.5 ug/ml) dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (2 ug/ml) dilution (Magenta). The Nuclear counterstain was DAPI (Blue).
-ve control 1: Anti-MIRO2 antibody [EPR29118-85] ab320739 at 1/50 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) antibody at 1/1000 (2 ug/ml) dilution.
-ve control 2: anti-COX IV mouse monoclonal antibody at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 ug/ml) dilution.
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