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Orbital shaker

轨道振动器

Company: Wisd Laboratory Instruments
Catalog#: SHO-1D
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DNA Slot Blot Repair Assay
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Date:
2015-04-20
[Abstract]  Ultraviolet (UV) irradiation induces helix distorting photolesions such as cyclobutane pyrimidine dimers (CPD) and pyrimidine-pyrimidone (6-4) photoproducts (6-4PP) which threaten genomic integrity if unrepaired. In mammals, nucleotide excision repair (NER) is the only pathway that removes UV-induced DNA damages. Here we describe DNA slot blot repair assay for quantitative detection of NER activity using DNA damage specific antibodies such as anti-CPD and anti-6-4PP. Briefly, genomic DNA irradiated with UV was isolated from cells, and the genomic DNA was vacuum-transferred to a nitrocellulose membrane using a Bio-Dot SF microfiltration apparatus (Bio-Rad). A monoclonal antibody that recognizes CPD or 6-4PP was applied to detect the remaining amount of photolesions in the genomic DNA. For ... [摘要]  紫外线(UV)照射诱导螺旋扭曲光致损伤,例如环丁烷嘧啶二聚体(CPD)和嘧啶 - 嘧啶酮(6-4)光产物(6-4PP),如果未修复则威胁基因组完整性。 在哺乳动物中,核苷酸切除修复(NER)是去除UV诱导的DNA损伤的唯一途径。 在这里我们描述DNA狭缝印迹修复测定NER活性使用DNA损伤特异性抗体如抗CPD和抗6-4PP的定量检测。 简言之,从细胞中分离用UV照射的基因组DNA,使用Bio-Dot SF微量过滤装置(Bio-Rad)将基因组DNA真空转移到硝酸纤维素膜上。 应用识别CPD或6-4PP的单克隆抗体来检测基因组DNA中残留的光损伤量。 对于均匀负载的上样控制,可以通过SYBR金染色进一步分析DNA在膜上。

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