Cell Line Information
● Cell line name: CEM/C1 (human acute lymphoblastic leukemia cells)
● Mutant gene: TP53(C242S)
● Genotype: Homozygous
● Species: Human
● Cell morphology: Suspension cells
● Complete medium composition: 1640 + 10% FBS + 1% P/S
● Passage ratio: 1:4–1:6
● Passage frequency: Passaged once every 2–3 days.
● Mycoplasma test: Negative

Gene Editing Information
● Construction method: The theoretical basis for constructing cellular point‑mutation models is homologous recombination repair. When the double strands of genomic DNA are cleaved by external factors, the cell’s DNA repair mechanisms are activated to repair the broken DNA ends; depending on the repair pathway, this can be classified as homologous recombination repair (HR) or non‑homologous end joining (NHEJ). Homologous recombination involves using a homologous DNA sequence as a template to achieve precise repair, whereas non‑homologous end joining entails the direct ligation of the broken DNA ends by cellular proteins, which may randomly insert or delete a variable number of nucleotides. Based on this theoretical framework, the construction of point‑mutation cell lines proceeds in three steps:

1. Site-specific DNA cleavage ： In this process, the Cas9 protein is transfected into the target cells together with the sgRNA. Under the guidance of the sgRNA, the Cas9 protein cleaves the DNA at the target site, generating a double-strand break.
2. Homologous recombination repair of DNA ： Following the induction of double-strand breaks in genomic DNA, an additional homologous repair template must be provided to the cell to enable repair via homologous recombination and to introduce a targeted mutation. This homologous template sequence is delivered into the cell by co-transfection with the sgRNA and Cas9 protein.
3. Positive cell screening ： For the point‑mutant cells generated by the aforementioned method, no additional reporter or resistance genes are introduced, thereby minimizing any off‑target effects on unedited genomic loci. Single‑clone cell lines bearing the intended point mutations are isolated from the edited cell pool and subjected to deep sequencing.

● Application scenarios: Liman Bio’s point‑mutation cell lines precisely mimic single‑base substitutions, making them ideal for elucidating gene function and investigating disease mechanisms. These platforms enable the generation of pathological cell models—such as those for cancer and genetic disorders—enabling validation of drug targets and assessment of drug resistance, thereby providing a standardized, highly reliable in vitro experimental system for basic research and new‑drug development.

● Validation data:

Cell Culture Instructions

● Cell recovery

1) Remove the cryovial from the liquid nitrogen tank or the −80°C freezer, and promptly place it in a 37°C water bath, gently swirling it to facilitate rapid thawing. To minimize the risk of contamination, you may first transfer the cryovial into a clean, disposable PE glove before immersing it in the water bath.

2) Once the contents of the cryovial have thawed, remove it immediately and wipe the exterior of the cryovial with a 75% ethanol‑soaked gauze.

3) Place the cryovial in a biosafety cabinet, transfer the contents of the cryovial into a centrifuge tube containing approximately 9 mL of complete culture medium, centrifuge at 300 × g for 5 minutes, and discard the supernatant. Discard the supernatant.

4) Resuspend the cells in an appropriate volume of complete culture medium, transfer them to a new culture flask, and incubate them in a 5% CO2, 37°C incubator under static conditions.

● Cell passage

Subculture of suspension cells can be performed by either replenishing with fresh medium or replacing the existing medium; the specific procedure is as follows:

1) Take a small aliquot of the cell suspension for cell counting and viability assessment; when the cell density reaches 1.5× 10⁶ When the cell density reaches cells/mL, perform subculture.

2) Add an adequate number of cells to a culture flask containing fresh culture medium, maintaining the cell density at 7 × 10^5 cells/mL.

3) Place the culture flask upright in a 5% CO2, 37°C incubator and incubate. If the cells reach the density required for passaging, proceed with subculturing.

4) Media change: Every 2–3 days. (Note: Use culture flasks with filter‑cap lids to maintain the balance of air and CO₂ in the medium.)

● Cell cryopreservation

After centrifuging to collect the cells, add an appropriate volume of cryopreservation solution (so that the cryopreserved cell density in each tube reaches…). 10⁶ ), place the cryovials into a cryovial box and store them at −80°C; after 24 hours, transfer the cryovials to liquid nitrogen for long-term storage.

Click the link below to view detailed information.

Product Manual | Grainman Easy Point Mutation Cell Line Construction Custom Services and Products

Service Guide | Q&A on LIMAN Point-Mutant Cell Line Construction Services [Version 2411]

Service Guide | Comprehensive Q&A on Site‑Directed Mutagenesis–Based Stable Cell Line Construction: From Concepts to Experimental Details—All in One Article! [Version 2512]