Catalogue No.
14040205
H129-2
H129-2
Cell Line Name
H129-2
Cell Line Description
Haploid embryonic mouse stem cell line established from inner cell mass derived from activated unfertilised oocytes from superovulated 129/Sv hybrid female mice. Teratoma formation in immune-compromised mice.
General Info
Species
Mouse
Links
Cellosaurus: CVCL_5G06
Release Conditions
Restricted - commercial organisations are required to complete the 'Cell Line Release Authorisation for Research Use in Commercial Organisations' release conditions form in the supporting documents section.
Characteristics
Receptors
Not known
Products
Haploid cells maintain an intact haploid genome without amplification or losses and show a wide differentiation potential. They can be used as a forward genetic screening tool.
Tissue of Origin
Embryo
Morphology
Small cells growing in large dense round colonies
Karyotype
Mixed (haploid and diploid), it is recommended that a haploid population of cells is isolated by fluorescence-activated cell sorting (FACs) after establishment of a growing culture.
Applications
Haploid cells can be used for gene identification by forward genetic screening, generation of homozygous mutation libraries, introduction of mutations into mice.
Disease
None Stated
Culture Conditions
Cell Type
Stem cell
Subculture Routine
At sub-confluence incubate using Accutase solution for 5 minutes at 37°C. Add 10 ml DMEM/F12 with 3 ml 7.5% BSA per 500 ml and pipette up and down 5 times to make a single cell suspension. Spin at 300 x g for 5 minutes and re-suspend in fresh 2i medium and plate into new tissue culture flask (at a ratio of 1:5 to 1:10). Establish new cultures on mitotically-arrested mouse embryonic fibroblast monolayers or gelatin coated cell culture plastic as above or at 4-5 x 10⁴ cells/cm². A complete media change must be performed at least every other day and cells subcultured every 2-3 days. Cultures must be kept at a low density with small colony sizes. Growth to high density or large colony size or prolonged subcultures will result in loss of haploid phenotype and increase in number of diploid cells as well as differentiation. Cultures must be incubated in a humidified 5% CO₂/95% air incubator at 37°C. Vials contain > 10⁶ cells. Contents can be thawed and placed in a 25 cm² flask. Please note: The haploid mouse stem cells upon resuscitation will have a viability of ~80% and a count of 2 x 10⁶ cells per ampoule. Upon cultivation with recommended culture conditions in a 25cm² flask H129-2 cells will attain a viability of ~95% and a cell yield of approximately 1 x 10⁶ cells.
Culture Medium
For 100ml of (2i) growth medium use: 100 ml NDiff 227 (Takara Clontech, catalogue number Y40002); 30 µl CHIR99021 (10 mM stock, Reagents Direct catalogue number 27-H76, 2mg); 10 µl PD0325901 (10 mM stock, store at -20°C; Reagents Direct catalogue number 39-C68, 2mg); 100,000 units LIF (=1 µg; eBioscience catalogue number 14-8521-80, mouse recombinant LIF, 25 µg); 5 ml 7.5% BSA (Life Technologies catalogue number 15260-037)
Growth Mode
Adherent - use gelatin (porcine)-coated tissue culture plastic. Feeder layers recommended for optimal growth (see H129-2 - Instructions for Culturing in the Product Documents section below)
Additional Info
Depositor
Dr Martin Leeb, Centre for Stem Cell Research, Wellcome Trust and Medical Research Council, Stem Cell Institute, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QR
Country of Origin
United Kingdom
Hazard Group (ACDP)
Hazard Group (ACDP) 2
Applications
References
Leeb, M. and Wutz, A. (2011). Derivation of haploid embryonic stem cells from mouse embryos. Nature 479, 131-134. PMID: 21900896
Bibliography
Leeb M, Walker R, Mansfield B, Nichols J, Smith A, Wutz A. (2012) Germline potential of parthenogenetic haploid mouse embryonic stem cells. Development, Sep;139(18):3301-5. doi: 10.1242/dev.083675. PMID: 22912412.Leeb M, Perry AC, Wutz A. (2015) Establishment and Use of Mouse Haploid ES Cells. Curr Protoc Mouse Biol., Jun 1;5(2):155-85. doi: 10.1002/9780470942390.mo140214.PMID:26069082
Documents
Images
Available Formats
- Frozen
- DNA-5µg (100ng/µl)
Citation Guidance
If use of this culture results in a scientific publication, it should be cited in the publication as: H129-2 (ECACC 14040205).
Biosafety Information
Unless specified otherwise, at ECACC we routinely handle all of our cell lines at containment level 2 in accordance with the ACDP guidelines (Advisory Committee on Dangerous Pathogens) (UK). All cell cultures have the potential to carry as yet unidentified adventitious agents. It is the responsibility of the end user to ensure that their facilities comply with biosafety regulations for their own country. ACDP Guidance: Biological agents: Managing the risks in laboratories and healthcare premises.
Further Information
The Culture Collections represent deposits of cultures from world-wide sources. While every effort is made to ensure details distributed by Culture Collections are accurate, Culture Collections cannot be held responsible for any inaccuracies in the data supplied. References where quoted are mainly attributed to the establishment of the cell culture and not for any specific property of the cell line, therefore further references should be obtained regarding cell culture characteristics. Passage numbers where given act only as a guide and Culture Collections does not guarantee the passage number stated will be the passage number received by the customer.
Cultures supplied by Culture Collections are for research purposes only. Enquiries regarding the commercial use of a cell line are referred to the depositor of the cell line. Some cell lines have additional special release conditions such as the requirement for a material transfer agreement to be completed by the potential recipient prior to the supply of the cell line. Please view the Terms & Conditions of Supply for more information.