Siglec-G Knockout RAW264.7 Cell Line

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LM02042030468

Product ID： LM02042030468

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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: Siglec-G Knockout RAW264.7 Cell Line
- Commodity ID: LM02042030468
- Gene Symbol: Siglec10 Siglecg
- NCBI Gene ID: 243958
- Ensembl ID: ENSMUSG00000030468
- Uniprot ID: Q80ZE3
- 宿主细胞 / 类型: RAW264.7/mouse mononuclear macrophage leukemia cells
- 规格: 1×10^6 cells/ cryovials
- 生长培养基: DMEM + 10% FBS + 1% P,S
- 筛选标记: N/A
- 生长特性: Adherent cells, irregular shape
- 培养条件: 37℃, 5% CO2 incubator, passage at a ratio of 1:6 to 1:8
- 倍增时间: ~12-30 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: A putative adhesion molecule that mediates sialic-acid-dependent binding to cells. It preferentially binds to alpha-2,3- or alpha-2,6-linked sialic acid. The sialic acid recognition site may be masked by cis interactions with sialic acids on the same cell surface. In the immune response, it seems to act as an inhibitory receptor upon ligand-induced tyrosine phosphorylation by recruiting cytoplasmic phosphatase(s) via their SH2 domain(s) that block signal transduction through dephosphorylation of signaling molecules. It is involved in the negative regulation of B-cell antigen receptor signaling and specifically acts on B1 cells to inhibit Ca2+ signaling, cellular expansion, and antibody secretion. The inhibition of B cell activation is dependent on PTPN6/SHP-1. In association with CD24, it may be involved in the selective suppression of the immune response to danger-associated molecular patterns (DAMPs) such as HMGB1, HSP70, and HSP90. In association with CD24, it may regulate the immune response of natural killer (NK) cells. It plays a role in the control of autoimmunity. During the initiation of adaptive immune responses by CD8-alpha+ dendritic cells, it inhibits cross-presentation by impairing the formation of MHC class I-peptide complexes. Its function seems to implicate the recruitment of PTPN6/SHP-1, which dephosphorylates NCF1 of the NADPH oxidase complex, consequently promoting phagosomal acidification.
- 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:
- Siglec10 Siglecg
01. Pre-warm the complete 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 cap and gently transfer the cell suspension to a sterile centrifuge tube containing 9mL of complete 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 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 1/6 to 1/8, 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 terminate 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.
02. Ensure that the cells to be frozen meet the freezing requirements. Check the following status under a microscope: healthy appearance and morphological characteristics, growth cycle (late logarithmic phase), no signs of contamination or senescence.
03. Digest and centrifuge the cells (refer to the subculture procedure for specific steps)
04. Add freezing solution to resuspend the cells at 1 mL per tube. After mixing evenly, dispense into cryovials.
05. Place the cells in a programmable freezing container and freeze in a -80℃ refrigerator.
06. Subsequently, transfer the cells to a liquid nitrogen tank for long-term storage.
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