KC-4110

CHOK1-IL7Ra-CRLF2-Low Cell Line

41829

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Background of CHOK1-IL7Ra-CRLF2-Low Cell Line

Interleukin-7 receptor α chain (IL-7Rα) single nucleotide polymorphisms (SNPs) are associated with susceptibility to immunopathologies like autoimmune and inflammatory diseases. The current hypothesis about underlying mechanisms is based on the regulation of IL-7 availability for self-reactive T cells by influencing the generation of a soluble (s)IL-7Rα variant.
CRLF2 overexpression has been described as a biomarker of poor prognosis in T-cell acute lymphoblastic leukemia (T-ALL).This gene encodes a member of the type I cytokine receptor family. The encoded protein is a receptor for thymic stromal lymphopoietin (TSLP). Together with the interleukin 7 receptor (IL7R), the encoded protein and TSLP activate STAT3, STAT5, and JAK2 pathways, which control processes such as cell proliferation and development of the hematopoietic system. Rearrangement of this gene with immunoglobulin heavy chain gene (IGH) on chromosome 14, or with P2Y purinoceptor 8 gene (P2RY8) on the same X or Y chromosomes is associated with B-progenitor acute lymphoblastic leukemia (ALL) and Down syndrome ALL.

Specifications

Catalog Number	KC-4110
Cell Line Name	CHOK1-IL7Ra-CRLF2-Low Cell Line
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous human IL7Ra and CRLF2 gene in low level
Quantity	One vial of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640 + 10% FBS + 10μg/ml Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/ml
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

CHOK1 IL7Ra-CRLF2-Low Cell Line was generated using a lentiviral vector expressing the human IL7Ra and CRLF2 sequence.

Characterization

Figure 1: Characterization of human IL7Ra and CRLF2 overexpression in the CHO-K1 human IL7Ra and CRLF2 stable clone using PCR sequence

Figure 2: Characterization of human IL7Ra and CRLF2 overexpression in the CHO-K1 human IL7Ra and CRLF2 stable clone using FACS.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/mL Puromycin)in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO₂ incubator.
8. Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage.

References

1. Lundtoft C, Seyfarth J, Jacobsen M. IL7RA genetic variants differentially affect IL-7Rα expression and alternative splicing: a role in autoimmune and infectious diseases? Genes Immun. 2020 Feb;21(2):83-90.
2.Maciel ALT, Wolch K, Emerenciano M, Mansur MB. CRLF2 overexpression defines an immature-like subgroup which is rescued through restoration of the PRC2 function in T-cell precursor acute lymphoblastic leukemia. Genes Chromosomes Cancer. 2022 Jul;61(7):437-442.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.