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HK EGFP-LaminA/H2B-mCherry
Tissue
子宫颈
Disease
癌症
Synonyms
HeLa Kyoto EGFP-LaminA and H2B-mCherry

Documents

HK EGFP-LaminA/H2B-mCherry

Cell line · Product number 300921

The HK EGFP-LaminA/H2B-mCherry cell line is a genetically engineered Hela Kyoto derived cell model developed to facilitate advanced studies in nuclear dyn...

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01General information

Description

HK EGFP-LaminA/H2B-mCherry 细胞系是一种基因工程改造的 Hela Kyoto 衍生细胞模型,旨在促进活细胞内核动力学和染色质组织的高级研究。该细胞系表达两种融合蛋白:EGFP(增强型绿色荧光蛋白)与层粘连蛋白 A 融合,mCherry(红色荧光蛋白)与组蛋白 H2B 融合。EGFP-Lamin A融合蛋白能突出核包膜,可观察细胞周期进展过程中或各种实验条件下核结构的变化。与此同时,H2B-mCherry 融合蛋白与 DNA 结合,发出鲜艳的红色荧光,标记染色质,从而能够实时观察有丝分裂和间期的染色体过程。

这些细胞对于实时成像应用非常宝贵,包括研究核完整性、DNA 复制和细胞衰老,以及研究核结构被破坏的疾病,如癌症和板层病。这种细胞系的双色荧光功能允许同时观察核膜和染色质,有助于全面了解核-胞质相互作用和染色质的时空组织。这种功能使其成为分子生物学研究和细胞生物物理学的重要工具,为基因表达调控机制、核组织和细胞周期提供了洞察力。

Tissue
子宫颈
Disease
癌症
Synonyms
  • HeLa Kyoto EGFP-LaminA and H2B-mCherry

02Classification

Organism
Human
Sex
Female
Growth properties
Adherent
GMO status
GMO-S1
GMO status details
GMO-S1: This HeLa Kyoto line contains EGFP-Lamin A and H2B-mCherry constructs enabling dual-color imaging of nuclear lamina and chromatin. This classification applies only within Germany and may differ elsewhere.
Biosafety level
1

03Characteristics

Morphology
具有镶嵌石形状的上皮样细胞

04Identifiers

Citation
HK EGFP-LaminA/H2B-mCherry(Cytion 目录号 300921)
NCBI TaxID
9606
Cellosaurus
CVCL_1D62
Depositor
艾伦伯格实验室(EMBL)

05Expression and susceptibility

Protein expression
EGFP-LaminA/H2B-mCherry
Products
组蛋白 H2B

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
Dissociation reagent
Accutase
Subculturing
去除粘附细胞上的旧培养基,用不含钙和镁的 PBS 冲洗。T25 烧瓶用 3-5 毫升 PBS,T75 烧瓶用 5-10 毫升。然后用 Accutase 完全覆盖细胞,T25 烧瓶用 1-2 毫升,T75 烧瓶用 2.5 毫升。让细胞在室温下孵育 8-10 分钟,使其分离。孵育后,用 10 毫升培养基轻轻混合细胞使其重悬,然后用 300xg 离心 3 分钟。弃去上清液,用新鲜培养基重悬细胞,并将其转移到已装有新鲜培养基的新烧瓶中。
Seeding density
1 × 10⁴ 个细胞/平方厘米
Fluid renewal
每周 2 至 3 次
Post-thaw recovery
解冻后,将细胞以5×10⁴个细胞/平方厘米的密度接种于培养皿中,使细胞从冷冻过程中恢复并至少贴壁24小时。
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

Donor age
30年
Ethnicity
非洲裔美国人