人PARP1 knockout HEK-293T cell line (ab266598)
概述
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产品名称
人PARP1 knockout HEK-293T cell line
参阅全部 PARP1 细胞裂解液 -
Parental Cell Line
HEK293T -
Organism
Human -
Mutation description
Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2 -
Passage number
<20 -
Knockout validation
Sanger Sequencing, Western Blot (WB) -
经测试应用
适用于: WBmore details -
Biosafety level
2 -
常规说明
Recommended control: Human wild-type HEK293T cell line (ab255449). 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, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and patents please refer to our limited use license and patent pages.
We will provide viable cells that proliferate on revival.
性能
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Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Kidney -
Cell type
epithelial -
STR Analysis
Amelogenin X D5S818: 8, 9 D13S317: 12, 14 D7S820: 11 D16S539: 9, 13 vWA: 16, 19 TH01: 7, 9.3 TPOX: 11 CSF1PO: 11, 12 -
Antibiotic resistance
Puromycin 1.00µg/ml -
Mycoplasma free
Yes -
存放说明
Shipped on Dry Ice. Store in liquid nitrogen. -
存储溶液
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
研究领域
靶标
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功能
Involved in the base excision repair (BER) pathway, by catalyzing the poly(ADP-ribosyl)ation of a limited number of acceptor proteins involved in chromatin architecture and in DNA metabolism. This modification follows DNA damages and appears as an obligatory step in a detection/signaling pathway leading to the reparation of DNA strand breaks. Mediates the poly(ADP-ribosyl)ation of APLF and CHFR. Positively regulates the transcription of MTUS1 and negatively regulates the transcription of MTUS2/TIP150. -
序列相似性
Contains 1 BRCT domain.
Contains 1 PARP alpha-helical domain.
Contains 1 PARP catalytic domain.
Contains 2 PARP-type zinc fingers. -
翻译后修饰
Phosphorylated by PRKDC. Phosphorylated upon DNA damage, probably by ATM or ATR.
Poly-ADP-ribosylated by PARP2. Poly-ADP-ribosylation mediates the recruitment of CHD1L to DNA damage sites.
S-nitrosylated, leading to inhibit transcription regulation activity. -
细胞定位
Nucleus. - Information by UniProt
相关产品
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KO cell lysates
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Related Products
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab266598于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 113 kDa.
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说明 |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 113 kDa. |
图片
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Sequencing chromatogram displaying sequence edit in exon 2
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All lanes : Anti-PARP1 antibody [E102] (ab32138) at 1/1000 dilution
Lane 1 : Wild-type HEK-293T cell lysate
Lane 2 : PARP1 knockout HEK-293T cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 113 kDa
Observed band size: 113 kDaLanes 1- 2: Merged signal (red and green). Green - ab32138 observed at 113 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab32138 was shown to react with PARP1 in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266598 (knockout cell lysate ab257017) was used. Wild-type HEK-293T and PARP1 knockout HEK-293T cell lysates were subjected to SDS-PAGE. ab32138 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 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.
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Homozygous: 1 bp insertion in exon 2
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Representative images of PARP1 knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.
实验方案
数据表及文件
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SDS download
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Datasheet download
文献 (0)
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