Mouse LRRK2 (N-terminal Apex2) LG knock-in SV40 MEF cell line available to order.
Recommended control: Mouse wild-type SV40 MEF cell line (ab283357).
Images
Key facts
- Cell type
- SV40 MEF
- Species or organism
- Mouse
- Tissue
- Embryo
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Homozygote. Large knock-in of APEX inserted in N-terminal LRKK2 gene
Mouse LRRK2 (N-terminal Apex2) LG knock-in SV40 MEF cell line available to order.
Recommended control: Mouse wild-type SV40 MEF cell line (ab283357).
Key facts
- Cell type
- SV40 MEF
- Species or organism
- Mouse
- Tissue
- Embryo
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Homozygote. Large knock-in of APEX inserted in N-terminal LRKK2 gene
- Concentration
Properties
- Gene name
- Lrrk2
- Gene editing type
- Knock-in
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing
Cell culture
- Biosafety level
- EU: 2 US: 2
- Adherent/suspension
- Adherent
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
- EMEM + 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 duration
- A few days
- Appropriate short-term storage conditions
- -80°C
- Appropriate long-term storage conditions
- -196°C
Notes
Recommended control: Mouse wild-type SV40 MEF cell line (ab283357). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: EMEM + 10% FBS
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.
- 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. Based on cell count, seed cells in an appropriate cell culture flask at a density of 2x104cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules.
4. Incubate the culture at 37°C incubator with 5% CO2. 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.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
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
The protein LRRK2 also known as leucine-rich repeat kinase 2 or dardarin is an enzyme with a molecular weight of approximately 286 kDa. It functions as a kinase meaning it adds phosphate groups to other proteins which affects their activity. LRRK2 is expressed in various tissues but it is highly abundant in the brain especially in regions such as the striatum and cortex. It has a significant role in cellular signaling processes due to its phosphorylation activity.
- Biological function summary
LRRK2 interacts with cellular mechanisms by regulating cytoskeletal dynamics autophagy and vesicle trafficking. It is a part of a larger complex that includes other proteins involved in these processes. The kinase activity of LRRK2 plays an essential part in maintaining neuronal health and function. It influences the process of autophagy which is a way cells clean themselves by removing damaged components and recycling them.
- Pathways
The action of LRRK2 is central to the mitogen-activated protein kinase (MAPK) and the mammalian target of rapamycin (mTOR) pathways. In these pathways LRRK2 interacts with other proteins such as mTOR and RPS6KB1. It modulates cellular processes like growth proliferation and response to stressors. Its kinase activity affects the phosphorylation state of targets within the pathways hence influencing biological outcomes like survival and apoptosis.
- Associated diseases and disorders
LRRK2 mutations have a significant connection to Parkinson's disease and Crohn's disease. In Parkinson's disease mutated LRRK2 leads to abnormal protein aggregation linking to proteins such as alpha-synuclein. For Crohn's disease LRRK2 influences the immune response and intestinal inflammation. These connections highlight LRRK2's role in the pathogenesis and contribute to understanding these complex disorders.
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1 product image
Sanger Sequencing - Mouse LRRK2 (N-terminal Apex2) LG knock-in SV40 MEF cell line (ab283306)
Homozygote. Large knock-in of APEX inserted in N-terminal LRKK2 gene
5’HA ---highlighted in Yellow
Lrrk2 5’UTR--- in dark red
3XFLAG--- in purple
HA---in blue
APEX2 --- Highlighted in light green
Lrrk2 Exon 1--- Highlighted in Dark green
Lrrk2 Exon 2--- Highlighted in Dark blue
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