8-氧代鸟嘌呤 DNA 糖基酶（Fpg） |NEB酶试剂 New England Biolabs

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产品信息

Fpg（甲酰胺嘧啶 [fapy]-DNA 糖基酶），也称作 8-氧代鸟嘌呤 DNA 糖基酶，既有 N-糖基酶活性也有 AP-裂解酶活性。N-糖基酶活性可以切下双链 DNA 上受损的嘌呤碱基，产生一个脱嘌呤（AP）位点。AP-裂解酶活性可以切割 AP 位点的 3´ 端和 5´ 端，从而去除 AP 位点，产生一个具有 3´ 和 5´ 磷酸的碱基缺口。被 Fpg 识别并切除的受损碱基包括：7,8-二羟基-8-氧代鸟嘌呤（8-氧代鸟嘌呤）、8-氧代腺嘌呤、fapy-鸟嘌呤、甲基-fapy-鸟嘌呤、fapy-腺嘌呤、黄曲霉毒素 B1-fapy-鸟嘌呤、5-羟基-胞嘧啶和 5-羟基尿嘧啶（1,2）。

产品来源

大肠杆菌菌株，携带 8-氧代鸟嘌呤 DNA 糖基酶（Fpg）基因（3）

产品类别:
DNA Repair Enzymes and Structure-specific Endonucleases Products

应用:
Polymerases for DNA Manipulation

  • 产品组分信息

    本产品提供以下试剂或组分：

    NEB # 名称 组分货号 储存温度 数量 浓度
    • M0240S     -20    
        Fpg M0240SVIAL -20 1 x 0.065 ml 8,000 units/ml
        NEBuffer™ 1 B7001SVIAL -20 1 x 1.25 ml 10 X
        Recombinant Albumin, Molecular Biology Grade B9200SVIAL -20 1 x 0.6 ml 20 mg/ml
    • M0240L     -20    
        Fpg M0240LVIAL -20 1 x 0.32 ml 8,000 units/ml
        NEBuffer™ 1 B7001SVIAL -20 1 x 1.25 ml 10 X
        Recombinant Albumin, Molecular Biology Grade B9200SVIAL -20 1 x 0.6 ml 20 mg/ml

  • 特性和用法

    单位定义

    1 单位指在 10 μl 反应体系中，37℃ 条件下，1 小时内能切割 10 pmol 含单个与胞嘧啶配对的 8-氧代鸟嘌呤的 34 bp 寡核苷酸双链所需要的酶量。

    反应条件

    1X NEBuffer™ 1
    Supplement with 100 µg/ml Recombinant Albumin, Molecular Biology Grade
    Incubate at 37°C

    1X NEBuffer™ 1
    10 mM Bis-Tris-Propane-HCl
    10 mM MgCl2
    1 mM DTT
    (pH 7 @ 25°C)

    彗星实验稀释

    彗星试验的推荐稀释倍数：1:103 到 1:104（4,5,6）。具体操作说明请登陆：http://cometassay.com。

    贮存溶液

    20 mM Tris-HCl
    50 mM NaCl
    0.5 mM EDTA
    200 µg/ml BSA
    50% Glycerol
    pH 8 @ 25°C

    热失活

    60°C for 10 minutes

    单位活性检测条件

    10 μl 总反应体系，含 1X NEBuffer 1、10 pmol 荧光标记寡核苷酸双链和 100 µg/ml BSA。

  • 相关产品

    单独销售的组分

    • NEBuffer 1
    • 重组白蛋白, 分子生物学级（不含动物成分）

  • 参考文献

    1. Tchou, J. et al. (1994). Substrate specificity of Fpg protein. J. Biol.Chem.. 269, 15318-15324.
    2. Hatahet, Z. et al. (1994). Newsubstrates for old enzymes. J. Biol.Chem.. 269, 18814-18820.
    3. Boiteux, S., O’Connor, T. and Laval, J. (1987). Formamidopyrimidine-DNA glycosylase of Escherichia coli: cloning and sequencing of the fpg structural gene and overproduction of the protein. EMBOJ.. 5, 18814-18820.
    4. Singh, N., McCoy, M., Tice, R. and Schneider, L. (1988). A simple technique for quantitation of low levels of DNA damage in individual cells. Exp.Cell Res.. 175, 184-191.
    5. Collins, A., Duthie, S. and Dobson, V. (1993). Direct enzymatic detection of endogenous oxidative base damage in human lymphocyte DNA. Carcinogenesis. 14, 1733-1735.
    6. Collins, A., Dusinska, M., Gedik, C. and Stetina, R. (1996). Oxidative damage to DNA: do we have a reliable biomarker?. Environ.Health Perspect.. 104, 465-469.
    7. Pflaum, M., Will, O., Mahler, H.-C. and Epe, B. (1998). DNA oxidation products determined with repair endonucleases in mammalian cells: types, basal levels and influence of cell proliferation. FreeRad. Res.. 29, 585-594.
    8. Hartwig, A., Dally, H. and Schlepegrell, R. (1996). Sensitive analysis of oxidative DNA damage in mammalian cells: use of the bacterial Fpg protein in combination with alkaline unwinding. Toxicol.Lett.. 88, 85-90.
    9. Czene, S. and Harms-Ringdahl, M. (1995). Detection of single strand breaks and formamidopyrimidine-DNA glycosylase-sensitive sites in DNA of cultured human fibroblasts. Mutat.Res.. 336, 235-242.

操作说明、说明书 & 用法

  • 操作说明

    1. Comet Assay – Modified for Detection of Oxidized Bases Using the Repair Endonucleases Fpg, hOGG1 and Endonuclease III (Nth)

  • 使用指南

    • DNA Damage and PreCR

工具 & 资源

  • 选择指南

    • Activities of DNA Repair Enzymes and Structure-specific Endonucleases
    • DNA Repair Enzymes on Damaged and Non-standard Bases
    • Properties of DNA Repair Enzymes and Structure-specific Endonucleases

FAQs & 问题解决指南

  • FAQs

    1. What is the activity of Fpg in the NEBuffers 1-4?
    2. Does Fpg cleave hydroxymethyl uracil?
    3. Does Fpg cut only one strand, or do they cause a double-strand break?
    4. What is the Molecular Weight of Fpg?
    5. What is the difference between using Fpg and hOOG1?
    6. Does the Fpg contain a tag?
    7. What buffer will work with both Endonuclease III and Fpg?
    8. What substrate is Fpg tested on?
    9. What is the activity of Fpg in rCutSmart Buffer?