CloneR™

Defined supplement for single-cell cloning of human ES and iPS cells
Catalog #
05888_C
Defined supplement for single-cell cloning of human ES and iPS cells
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Overview

CloneR™ is a defined, serum-free supplement designed to increase the cloning efficiency and single-cell survival of human embryonic stem cells (ES cells) and induced pluripotent stem cells (iPS cells). CloneR™ enables the robust generation of clonal cell lines without single-cell adaptation, thus minimizing the risk of acquiring genetic abnormalities.

CloneR™ is compatible with the TeSR™ family of media for human ES and iPS cell maintenance as well as your choice of cell culture matrix.

For improved single-cell survival in additional applications, and higher cloning efficiency, try our CloneR™2 supplement.
Advantages
• Greatly facilitates the process of genome editing of human ES and iPS cells
• Compatible with any TeSR™ maintenance medium and your choice of cell culture matrix
• Does not require adaptation to single-cell passaging
• Increases single-cell survival at clonal density across multiple human ES and iPS cell lines
Subtype
Supplements
Cell Type
Pluripotent Stem Cells
Species
Human
Application
Cell Culture
Brand
CloneR
Area of Interest
Cell Line Development, Disease Modeling, Stem Cell Biology
Formulation
Serum-Free

Scientific Resources

Product Documentation

Document Type Product Name Catalog # Lot # Language
Document Type
Product Information Sheet
Product Name
CloneR™
Catalog #
05888
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
CloneR™, 5 Pack
Catalog #
05889
Lot #
All
Language
English
Document Type
Safety Data Sheet
Product Name
CloneR™
Catalog #
05888
Lot #
All
Language
English

Educational Materials (25)

Brochure
Products for Human Pluripotent Stem Cells
Brochure
CloneR™ Supplement for the Increased Cloning Efficiency and Single-Cell Survival of hPSCs
Video
mTeSR™1: Standardized Medium for the Feeder-Independent Maintenance of hESCs & hiPSCs
1:27
mTeSR™1: Standardized Medium for the Feeder-Independent Maintenance of hESCs & hiPSCs
Video
Limitless Potential: Do More with TeSR™
3:10
Limitless Potential: Do More with TeSR™
Video
A Guide to Passaging Human Pluripotent Stem Cells Using mTeSR™1
5:58
A Guide to Passaging Human Pluripotent Stem Cells Using mTeSR™1
Video
How to Set Up an Assay with the hPSC Genetic Analysis Kit Experiment
8:33
How to Set Up an Assay with the hPSC Genetic Analysis Kit Experiment
Video
mTeSR™1: The Most Published, Feeder-Independent hESC & hiPSC Maintenance Medium
1:45
mTeSR™1: The Most Published, Feeder-Independent hESC & hiPSC Maintenance Medium
Webinar
Maintaining and Assessing High-Quality hPSC Cultures
55:53
Maintaining and Assessing High-Quality hPSC Cultures
Webinar
Considerations for High-Efficiency Genome Editing of Human Pluripotent Stem Cells
50:01
Considerations for High-Efficiency Genome Editing of Human Pluripotent Stem Cells
Webinar
Nature Research Round Table: Human Pluripotent Stem Cell Lines as Disease Models
17:30
Nature Research Round Table: Human Pluripotent Stem Cell Lines as Disease Models
Webinar
Nature Research Round Table: Best Practices for the QC of Genome-Edited hPSC Lines - Panel Discussion
31:25
Nature Research Round Table: Best Practices for the QC of Genome-Edited hPSC Lines - Panel Discussion
Webinar
Nature Research Round Table: Genome Editing in Human Pluripotent Stem Cells
17:45
Nature Research Round Table: Genome Editing in Human Pluripotent Stem Cells
Webinar
Re-Creating Disease with Kidney Organoids and CRISPR
1:14:36
Re-Creating Disease with Kidney Organoids and CRISPR
Webinar
Highly Efficient Single-Cell Human Pluripotent Stem Cell Cloning and Robust Cardiomyocyte Differentiation
48:54
Highly Efficient Single-Cell Human Pluripotent Stem Cell Cloning and Robust Cardiomyocyte Differentiation
Webinar
Modeling Alzheimer's risk using human TREM2-knockout microglia
56:09
Modeling Alzheimer's risk using human TREM2-knockout microglia
Webinar
Nature Research Round Table: hPSC Lines for Cell Therapies - Panel Discussion
29:03
Nature Research Round Table: hPSC Lines for Cell Therapies - Panel Discussion
Webinar
Nature Research Round Table: Retinal Cell Therapy Using Human Embryonic Stem Cells
19:35
Nature Research Round Table: Retinal Cell Therapy Using Human Embryonic Stem Cells
Webinar
Nature Research Round Table: The Process of Human Pluripotent Stem Cell Adaptation
18:48
Nature Research Round Table: The Process of Human Pluripotent Stem Cell Adaptation
Webinar
Development, Compatibility, and Applications of mTeSR™ Plus; an Enhanced Medium for the Maintenance of Human Pluripotent Stem Cells (hPSCs)
42:10
Development, Compatibility, and Applications of mTeSR™ Plus; an Enhanced Medium for the Maintenance of Human Pluripotent Stem Cells (hPSCs)
Webinar
Using CRISPR/Cas9 to Model Stem Cell Organization and Dynamics
71:57
Using CRISPR/Cas9 to Model Stem Cell Organization and Dynamics
Webinar
Facilitating hPSC Single Cell Seeding Workflows Using CloneR™2
12:49
Facilitating hPSC Single Cell Seeding Workflows Using CloneR™2
Webinar
Best Practice Criteria for Pluripotent Stem Cell Lines
55:52
Best Practice Criteria for Pluripotent Stem Cell Lines
Webinar
Nature Research Round Table: Parkinson's Disease Therapy with Human Embryonic Stem Cells
24:46
Nature Research Round Table: Parkinson's Disease Therapy with Human Embryonic Stem Cells
Webinar
Nature Research Round Table: Genomic Integrity of Human Pluripotent Stem Cells
15:47
Nature Research Round Table: Genomic Integrity of Human Pluripotent Stem Cells
Webinar
Nature Research Round Table: Identifying Acquired and Background Genetic Variants in Human Pluripotent Stem Cells
16:56
Nature Research Round Table: Identifying Acquired and Background Genetic Variants in Human Pluripotent Stem Cells
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Product Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Data and Publications

Data

hPSC Single-Cell Cloning Workflow with CloneR™

Figure 1. hPSC Single-Cell Cloning Workflow with CloneR™

On day 0, human pluripotent stem cells (hPSCs) are seeded as single cells at clonal density (e.g. 25 cells/cm2) or sorted at 1 cell per well in 96-well plates in TeSR™ (mTeSR™1 or TeSR™-E8™) medium supplemented with CloneR™. On day 2, the cells are fed with TeSR™ medium containing CloneR™ supplement. From day 4, cells are maintained in TeSR™ medium without CloneR™. Colonies are ready to be picked between days 10 - 14. Clonal cell lines can be maintained long-term in TeSR™ medium.

CloneR™ Increases the Cloning Efficiency of hPSCs and is Compatible with Multiple hPSC Lines and Seeding Protocols

Figure 2. CloneR™ Increases the Cloning Efficiency of hPSCs and is Compatible with Multiple hPSC Lines and Seeding Protocols

TeSR™ medium supplemented with CloneR™ increases hPSC cloning efficiency compared with cells plated in TeSR™ containing ROCK inhibitor. Cells were seeded (A) at clonal density (25 cells/cm2) in mTeSR™1 and TeSR™-E8™ and (B) by single-cell deposition using FACS (seeded at 1 cell/well) in mTeSR™1.

CloneR™ Increases the Cloning Efficiency of hPSCs at Low Seeding Densities

Figure 3. CloneR™ Increases the Cloning Efficiency of hPSCs at Low Seeding Densities

hPSCs plated in mTeSR™1 supplemented with CloneR™ demonstrated significantly increased cloning efficiencies compared to cells plated in mTeSR™1 containing ROCK inhibitor (10μM Y-27632). Shown are representative images of alkaline phosphatase-stained colonies at day 7 in individual wells of a 12-well plate. H1 human embryonic stem (hES) cells were seeded at clonal density (100 cells/well, 25 cells/cm2) in mTeSR™1 supplemented with (A) ROCK inhibitor or (B) CloneR™ on Vitronectin XF™ cell culture matrix.

CloneR™ Yields Larger Single-Cell Derived Colonies

Figure 4. CloneR™ Yields Larger Single-Cell Derived Colonies

hPSCs seeded in mTeSR™1 supplemented with CloneR™ result in larger colonies than cells seeded in mTeSR™1 containing ROCK inhibitor (10μM Y-27632). Shown are representative images of hPSC clones established after 7 days of culture in mTeSR™1 supplemented with (A) ROCK inhibitor or (B) CloneR™.

Clonal Cell Lines Established Using CloneR™ Display Characteristic hPSC Morphology

Figure 5. Clonal Cell Lines Established Using CloneR™ Display Characteristic hPSC Morphology

Clonal cell lines established using mTeSR™1 or TeSR™-E8™ medium supplemented with CloneR™ retain the prominent nucleoli and high nuclear-to-cytoplasmic ratio characteristic of hPSCs. Representative images at passage 7 after cloning are shown for clones derived from the parental (A) H1 hES cell and (B) WLS-1C human induced pluripotent stem (iPS) cell lines.

Clonal Cell Lines Established with CloneR™ Express High Levels of Undifferentiated Cell Markers

Figure 6. Clonal Cell Lines Established with CloneR™ Express High Levels of Undifferentiated Cell Markers

hPSC clonal cell lines established using mTeSR™1 supplemented with CloneR™ express comparable levels of undifferentiated cell markers, OCT4 (Catalog #60093) and TRA-1-60 (Catalog #60064), as the parental cell lines. (A) Clonal cell lines established from parental H1 hES cell line. (B) Clonal cell lines established from parental WLS-1C hiPS cell line. Data is presented between passages 5 - 7 after cloning and is shown as mean ± SEM; n = 2.

Clonal Cell Lines Established Using CloneR™ Display a Normal Karyotype

Figure 7. Clonal Cell Lines Established Using CloneR™ Display a Normal Karyotype

Representative karyograms of clones derived from parental (A) H1 hES cell and (B) WLS-1C hiPS cell lines demonstrate that the clonal lines established with CloneR™ have a normal karyotype. Cells were karyotyped 5 passages after cloning, with an overall passage number of 45 and 39, respectively.

Clonal Cell Lines Established Using CloneR™ Display Normal Growth Rates

Figure 8. Clonal Cell Lines Established Using CloneR™ Display Normal Growth Rates

Fold expansion of clonal cell lines display similar growth rates to parental cell lines. Shown are clones (red) and parental cell lines (gray) for (A) H1 hES cell and (B) WLS-1C hiPS cell lines.

Cell morphology images of ES cells plated in  mTeSR™1 and mTeSR™ Plus and supplemented with CloneR™ immediately following RNP electroporation.

Figure 9. Representative Cell Morphology 24 Hours After RNP Electroporation in mTeSR™1 and mTeSR™ Plus

H1-eGFP ES cells were plated in (A) mTeSR™1 and (B) mTeSR™ Plus and supplemented with CloneR™ immediately following RNP electroporation. Images were taken 24 hours after electroporation.

Cell images of human ES colonies plated in mTeSR™1 and mTeSR™ Plus and supplemented with CloneR™ on CellAdhere™ Vitronectin™ XF™-coated plates.

Figure 10. Clones Derived in mTeSR™ Plus are Larger and Ready to Be Picked at an Earlier Timepoint

Representative images of human ES (H9) colonies taken 8 days following singlecell plating at clonal density (25 cells/cm²) in either (A) mTeSR™1 or (B) mTeSR™ Plus supplemented with CloneR™ on CellAdhere™ Vitronectin™ XF™-coated plates.

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