Human CTNNB1 (beta Catenin) knockout HeLa cell line (ab255352)
製品の概要
-
製品名
Human CTNNB1 (beta Catenin) knockout HeLa cell line
beta Catenin ライゼート 製品一覧 -
Parental Cell Line
HeLa -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 3 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
アプリケーション
適用あり: WBmore details -
Biosafety level
2 -
特記事項
Recommended control: Human wild-type HeLa cell line (ab255448). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: DMEM (High Glucose) + 10% FBS
Initial handling guidelines: Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method. Based on cell count, seed cells in an appropriate cell culture flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules.
4. Incubate the culture at 37°C incubator with 5% CO2. Cultures should be monitored daily.Subculture guidelines:
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
We will provide viable cells that proliferate on revival.
製品の特性
-
Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Cervix -
Cell type
epithelial -
Disease
Adenocarcinoma -
Gender
Female -
STR Analysis
Amelogenin X D5S818: 11, 12 D13S317: 12, 13.3 D7S820: 8, 12 D16S539: 9, 10 vWA: 16, 18 TH01: 7 TPOX: 8, 12 CSF1PO: 9, 10 -
Mycoplasma free
Yes -
保存方法
Shipped on Dry Ice. Store in liquid nitrogen. -
バッファー
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
研究分野
ターゲット情報
-
機能
Key dowstream component of the canonical Wnt signaling pathway. In the absence of Wnt, forms a complex with AXIN1, AXIN2, APC, CSNK1A1 and GSK3B that promotes phosphorylation on N-terminal Ser and Thr residues and ubiquitination of CTNNB1 via BTRC and its subsequent degradation by the proteasome. In the presence of Wnt ligand, CTNNB1 is not ubiquitinated and accumulates in the nucleus, where it acts as a coactivator for transcription factors of the TCF/LEF family, leading to activate Wnt responsive genes.
Involved in the regulation of cell adhesion. The majority of beta-catenin is localized to the cell membrane and is part of E-cadherin/catenin adhesion complexes which are proposed to couple cadherins to the actin cytoskeleton. -
組織特異性
Expressed in several hair follicle cell types: basal and peripheral matrix cells, and cells of the outer and inner root sheaths. Expressed in colon. -
関連疾患
Defects in CTNNB1 are associated with colorectal cancer (CRC) [MIM:114500].
Note=Activating mutations in CTNNB1 have oncogenic activity resulting in tumor development. Somatic mutations are found in various tumor types, including colon cancers, ovarian and prostate carcinomas, hepatoblastoma (HB), hepatocellular carcinoma (HCC). HBs are malignant embryonal tumors mainly affecting young children in the first three years of life.
Defects in CTNNB1 are a cause of pilomatrixoma (PTR) [MIM:132600]; a common benign skin tumor.
Defects in CTNNB1 are a cause of medulloblastoma (MDB) [MIM:155255]. MDB is a malignant, invasive embryonal tumor of the cerebellum with a preferential manifestation in children.
Defects in CTNNB1 are a cause of susceptibility to ovarian cancer (OC) [MIM:167000]. Ovarian cancer common malignancy originating from ovarian tissue. Although many histologic types of ovarian neoplasms have been described, epithelial ovarian carcinoma is the most common form. Ovarian cancers are often asymptomatic and the recognized signs and symptoms, even of late-stage disease, are vague. Consequently, most patients are diagnosed with advanced disease.
Note=A chromosomal aberration involving CTNNB1 is found in salivary gland pleiomorphic adenomas, the most common benign epithelial tumors of the salivary gland. Translocation t(3;8)(p21;q12) with PLAG1. -
配列類似性
Belongs to the beta-catenin family.
Contains 12 ARM repeats. -
翻訳後修飾
Phosphorylation by GSK3B requires prior phosphorylation of Ser-45 by another kinase. Phosphorylation proceeds then from Thr-41 to Ser-37 and Ser-33.
EGF stimulates tyrosine phosphorylation. Phosphorylation on Tyr-654 decreases CDH1 binding and enhances TBP binding.
Ubiquitinated by the SCF(BTRC) E3 ligase complex when phosphorylated by GSK3B, leading to its degradation. Ubiquitinated by a E3 ubiquitin ligase complex containing UBE2D1, SIAH1, CACYBP/SIP, SKP1, APC and TBL1X, leading to its subsequent proteasomal degradation. -
細胞内局在
Cytoplasm. Nucleus. Cytoplasm > cytoskeleton. Cell junction > adherens junction. Cell junction. Cell membrane. Cytoplasmic when it is unstabilized (high level of phosphorylation) or bound to CDH1. Translocates to the nucleus when it is stabilized (low level of phosphorylation). Interaction with GLIS2 and MUC1 promotes nuclear translocation. Interaction with EMD inhibits nuclear localization. - Information by UniProt
関連製品
-
KO cell lysates
-
Related Products
- Anti-beta Catenin antibody [E247] - BSA and Azide free (ab196204)
- Anti-beta Catenin non-phospho(active) S37/T41 antibody [EPR23969-131] (ab246504)
- Anti-beta Catenin non-phospho(active) S37/T41 antibody [EPR23969-131] - BSA and Azide free (ab278503)
- Anti-beta Catenin antibody [E247] - ChIP Grade (ab32572)
- Anti-beta Catenin antibody [15B8] (ab6301)
アプリケーション
The Abpromise guarantee
Abpromise保証は、 次のテスト済みアプリケーションにおけるab255352の使用に適用されます
アプリケーションノートには、推奨の開始希釈率がありますが、適切な希釈率につきましてはご検討ください。
アプリケーション | Abreviews | 特記事項 |
---|---|---|
WB |
Use at an assay dependent concentration. Predicted molecular weight: 85 kDa.
|
特記事項 |
---|
WB
Use at an assay dependent concentration. Predicted molecular weight: 85 kDa. |
画像
-
All lanes : Anti-beta Catenin non-phospho(active) S37/T41 antibody [EPR23969-131] (ab246504) at 1/1000 dilution
Lane 1 : Wild-type HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : CTNNB1 (beta Catenin) knockout HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 800CW) (ab216773) and Goat Anti-Mouse IgG H&L (IRDye® 680RD) (ab216776) at 1/10000 dilution
Predicted band size: 85 kDa
Observed band size: 95 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBST
The molecular weight observed is consistent with what has been described in the literature (PMID: 16288032).
Lanes 1-2: Merged signal (red and green). Green - ab246504 observed at 92 kDa. Red - loading control ab8245 (Mouse monoclonal [6C5] to GAPDH) observed at 36 kDa.
ab246504 Anti-beta Catenin (non-phospho(active) S37/T41) antibody [EPR23969-131] was shown to specifically react with beta Catenin in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab255352 (knockout cell lysate ab263756) was used.
Wild-type and beta Catenin knockout samples were subjected to SDS-PAGE. ab246504 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.
An extra band around 45 kDa was observed.
-
All lanes : Anti-beta Catenin antibody [E247] - ChIP Grade (ab32572) at 1/500 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : CTNNB1 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 85 kDa
Observed band size: 86 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab32572 observed at 86 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab32572 was shown to react with CTNNB1 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255352 (knockout cell lysate ab263756) was used. Wild-type HeLa and CTNNB1 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32572 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 500 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
-
Homozygous: 1 bp deletion in exon 3.
プロトコール
データシートおよび資料
-
SDS download
-
Datasheet download
参考文献 (0)
ab255352 は論文での使用が確認できていません。