SOST Knockout MC3T3-E1 Cell Line

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LM02040001494

Product ID： LM02040001494

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产品描述
细胞复苏
细胞传代
细胞冻存
抗体验证结果

- Commodity name: SOST Knockout MC3T3-E1 Cell Line
- Commodity ID: LM02040001494
- Gene Symbol: Sost
- Ensembl ID: ENSMUSG00000001494
- Uniprot ID: Q99P68
- 宿主细胞 / 类型: MC3T3-E1/Mouse calvarial preosteoblast subclone 14
- NCBI Gene ID: 74499
- 规格: 1×10^6 cells/ cryovials
- 生长培养基: MEMα＋10% FBS＋1% P,S
- 筛选标记: N/A
- 生长特性: Adherent cells, fibroblast-like
- 培养条件: 37℃, 5% CO2 incubator, passage 1/3 to 1/4
- 倍增时间: 24-48 hours
- 参考换液频率: 2~3 times/week
- 支原体检测结果: Negative
- 敲除效率（Sanger测序）: 100%
- 蛋白质组验证结果: N/A
- 抗体货号: Adding
- 目标基因介绍: Negative regulator of bone growth that acts through inhibition of Wnt signaling and bone formation.
- 细胞开发路径: A stable KO cell pool was generated using the CRISPR-RNP method; Sanger sequencing results showed 100% knockout efficiency in the KO cell pool
- 应用: Highly efficient gene knockout cell lines (KO Cell Line) are particularly suitable for preliminary functional analysis, the development of complex disease models, precision drug screening, and a wide range of gene discovery research.
Key words:
- Sost
01. Pre-warm the complete culture medium in a 37℃ water bath.
02. Thaw the cryovials in a 37℃ water bath for 1-2 minutes.
03. Transfer the cryovials to a biosafety cabinet and wipe the surface with 70% ethanol.
04. Unscrew the cryovials, and gently transfer the cell suspension to a sterile centrifuge tube containing 9mL of complete culture medium.
05. Centrifuge at 125g for 5-7 minutes at room temperature, and discard the supernatant.
06. Resuspend the cell pellet with 5mL of complete culture medium, and transfer the cell suspension to a T25 culture flask.
07. Transfer the cells to a 37℃, 5% CO2 incubator for culture.
08. Reference passage ratio: passage 1/3 to 1/4, confluent in 2-3 days.
01. When the cell confluence in the culture bottle reaches 80%-90%, the cells can be passaged.
02. Take out the culture medium, PBS, trypsin (0.25% Trypsin_EDTA Gibco 25200-056), etc., from the 4℃ refrigerator and place them in a 37℃ water bath. When the temperature is close to 37℃, take them out, spray 75% alcohol on the surface of the bottle, and then place them in a biosafety cabinet.

03. Take out the culture bottle to be passaged from the incubator, spray 75% alcohol on the bottle, and then place it in a biosafety cabinet.
04. To avoid disrupting the cells, rinse the cells with PBS along the upper wall of the culture bottle. After washing the cells, discard the solution. Add 2mL to T25.
05. Add the corresponding volume of trypsin (1.5mL for T75, 0.5mL for T25), and gently shake the bottle to spread the trypsin evenly over the bottom of the cells. The amount can be appropriately increased or decreased according to the actual situation. After about 1-2 minutes, when most of the cells have detached, add the corresponding volume of complete culture medium to stop digestion, and gently blow the cells with a 5mL pipette until all the cells have detached.
06. Transfer the cell suspension to a 15mL centrifuge tube, centrifuge at 300g for 5min, and discard the supernatant.
07. Resuspend the cells with 5mL of complete culture medium, adjust the inoculation ratio as needed, and supplement the complete culture medium in the culture bottle. Add 13-15mL to T75, 5mL to T25, and add 1% double antibody.
08. Cover the bottle cap tightly, gently shake the bottle to mix the cells evenly, and then place it in a 37℃, 5% CO2 incubator.
01. Prepare freezing solution and pre-chill in advance.
02. Ensure that the cells to be cryopreserved meet the cryopreservation requirements. Check the following conditions under a microscope: healthy appearance and morphological characteristics, growth cycle (late logarithmic phase), no signs of contamination or degradation.
03. Digest and centrifuge the cells (refer to the subculture procedure for specific steps)
04. Add freezing solution to resuspend the cells according to the amount of 1mL per tube, mix well and aliquot into freezing tubes.
05. Place the cells in a programmed cooling box and freeze in a -80℃ refrigerator.
06. Subsequently, transfer the cells to a liquid nitrogen tank for long-term storage.
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