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NRK-EGFP2-Nup50
Tissue
肾脏
Synonyms
NRK EGFP2-Nup50

Documents

NRK-EGFP2-Nup50

Cell line · Product number 500726

The NRK-EGFP2-Nup50 cell line is a clonal stable cell line derived from normal rat kidney (NRK) cells. This cell line was generated through the transfecti...

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1

01General information

Description

NRK-EGFP2-Nup50细胞系是从正常大鼠肾脏(NRK)细胞中提取的克隆稳定细胞系。这种细胞系是通过转染含有编码增强型绿色荧光蛋白(EGFP)和核蛋白 50(Nup50)融合蛋白基因的环状质粒,然后进行耐药性选择而产生的。结果,约 50%的细胞表达 EGFP3-Nup50 融合蛋白,从而实现了 Nup50 在细胞环境中的可视化和追踪。

Nup50 是核孔复合体的重要组成部分,负责调节细胞核和细胞质之间的分子运输。EGFP3 标签可用于活细胞成像和其他基于荧光的技术,以研究 Nup50 的定位、动态和相互作用。尽管是一种稳定的细胞系,NRK-EGFP2-Nup50 细胞仍表现出一些变异,这表明细胞间 EGFP3-Nup50 融合蛋白的表达水平存在差异。

这种细胞系对于研究核胞质转运、核孔复合体动力学以及 Nup50 在各种细胞过程中的功能作用特别有价值。NRK-EGFP2-Nup50 细胞适用于一系列实验方法,包括光漂白后荧光恢复(FRAP)、荧光相关光谱(FCS)和其他先进的显微镜技术。这些研究可以让人们深入了解核转运的分子机制,有助于理解与核转运功能障碍相关的疾病,如某些癌症和神经退行性疾病。

Tissue
肾脏
Synonyms
  • NRK EGFP2-Nup50

02Classification

Organism
Rat
Growth properties
Adherent
Biosafety level
1

03Characteristics

Morphology
纺锤形纤维母细胞

04Identifiers

Citation
NRK-EGFP2-Nup50(Cytion 目录号 500726)
NCBI TaxID
10116
Cellosaurus
CVCL_AV93
Depositor
艾伦伯格实验室(EMBL)

05Expression and susceptibility

Receptors expressed
表皮生长因子(EGF),增殖刺激活性(MSA)
Protein expression
EGFP3-Nup50
Products
NUP50(核蛋白 50)

06Handling and culture

Culture medium
DMEM,w:4.5 克/升葡萄糖,w:4 毫摩尔 L-谷氨酰胺,w:3.7 克/升 NaHCO3,w:1.0 毫摩尔丙酮酸钠(Cytion 编号 820300a)。
Incubation atmosphere

37°C, 5%CO2, 加湿环境。

Supplements
在培养基中添加 10% FBS、0.5 mg/mL G418
Dissociation reagent
Accutase
Subculturing
丢弃旧培养基,用 PBS 冲洗细胞。加入新配制的 0.025%胰酶/0.02%乙二胺四乙酸溶液,加热至 37 摄氏度,等待细胞分离,通常需要 5 分钟左右。加入新鲜培养基中和胰蛋白酶,然后将细胞混合物转移到离心管中并离心。离心后,除去上清液,用新鲜培养基重悬细胞团,然后将悬浮液转移到新的烧瓶中。将 G418 加入培养基中,使其最终浓度达到 0.5 mg/ml
Seeding density
2至4×10⁴个细胞/平方厘米
Fluid renewal
每周 2 至 3 次
Thawing and culturing
  1. Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

  2. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

  3. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

  4. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

  5. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

  6. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

  7. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

  8. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

07Quality control and cryopreservation

Sterility

使用基于 PCR 的检测方法和基于发光的支原体检测方法排除支原体污染。

为确保没有细菌、真菌或酵母菌污染,每天都要对细胞培养物进行目视检查。

Freeze medium
我们使用完全生长培养基(包括 FBS)+10% DMSO 或 CM-1(Cytion 商品目录编号 800100)作为冷冻保存培养基,以获得足够的解冻后存活率,CM-1(Cytion 商品目录编号 800100)含有优化的渗透保护剂和代谢稳定剂,可提高恢复能力并减少冷冻引起的应激。
Storage conditions

如需长期保存,可将样品瓶放入气相液氮中,温度约为 -150 至 -196 ℃。在-80 °C下保存只能作为转入液氮前的短暂过渡。

Shipping conditions

冷冻保存的细胞系用干冰装运,并用经过验证的隔热包装和足够的制冷剂在整个运输过程中保持约 -78 °C 的温度。收到货后,应立即检查容器,并立即将小瓶转移到适当的储藏室。

08Donor

Breed / subspecies
奥斯本-门德尔