热搜词：
注册
登录
CN
  • CN
  • EN

产品中心 分子酶 分子诊断&NGS测序相关

所有图片（1/3）

T4 DNA Polymerase

DNA-directed DNA polymerase

价格 ￥760.00 （ 供应商现货 ： 3-5个工作日）
货号 UA070070
规格
数量
收藏 分享
产品介绍

产品介绍 评论(0)

产品规格

  • 分子别名

    DNA-directed DNA polymerase

  • 表达宿主

    E.coli

  • 分子量

    105kDa (Reducing)

  • 纯度

    ＞95% by SDS-PAGE and HPLC

  • 活性

    3 U/μl

  • 标记

    Unconjugated

  • 标签

    His Tag

  • 性状

    Liquid

  • 缓冲体系

    100 mM K3PO4, 1 mM DTT, 50% Glycerol, pH 6.5 @ 25°C

  • 储存条件

    Store at -25 ~ -15℃ for 2 years

  • 文献引用

    1. Capson TL, Peliska JA, Kaboord BF, Frey MW, Lively C, Dahlberg M, Benkovic SJ. Kinetic characterization of the polymerase and exonuclease activities of the gene 43 protein of bacteriophage T4. Biochemistry. 1992 Nov 17;31(45):10984-94.
    2. Tanguy Le Gac N, Delagoutte E, Germain M, Villani G. Inactivation of the 3'-5' exonuclease of the replicative T4 DNA polymerase allows translesion DNA synthesis at an abasic site. J Mol Biol. 2004 Mar 5;336(5):1023-34.

背景介绍

  • T4 DNA polymerase catalyzes the synthesis of DNA in the 5´→3´ direction and requires the presence of template and primer. This enzyme possesses 3´→5´ exonuclease activity, which is much higher than that found in DNA polymerase I (E. coli). Unlike E. coli DNA polymerase I, T4 DNA polymerase lacks 5’ →3’ exonuclease activity. T4 DNA polymerase can be used to Removal of 3’ overhangs or fill-in of 5’ overhangs to form blunt ends.

产品组分

  • 3 U/μl T4 DNA Polymerase, 100 mM K3PO4, 1 mM DTT, 50% Glycerol, pH 6.5 @ 25°C
    10* Reaction Buffer: 500 mM NaCl, 100 mM Tris-HCl, 100 mM MgCl2, 1000 µg/ml Recombinant Albumin, (pH 7.9 @ 25°C)

操作步骤


  • Incubate the following reaction at 12°C for 15 minutes

    Component

    Final Concentration

    10X Reaction Buffer

    1X

    dNTP Mix (10 mM)

    100 mM each

    T4 DNA Polymerase (3,000 U/ml)

    1000 U/ml

    DNA

    10 ug

    Nuclease-free Water

    to 10 µl

    Total Reaction Volume

    10 µl

    Stop reaction by adding EDTA to a final concentration of 10 mM and heating to 75°C for 20 minutes

注意事项

  • Elevated temperatures, excessive amounts of enzyme, failure to supplement with dNTPs or long reaction times will result in recessed ends due to the 3´ → 5´ exonuclease activity of the enzyme.

酶活定义
  • One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid insoluble material in 30 minutes at 37°C

  • 生物活性

    • The experiment was designed to synthesize two complementary DNA Oligos. Small DNA fragments with 5 'sticky ends were synthesized by annealing, and the sticky ends were completed by T4 DNA Polymerase to form flat ends. 20% Native PAGE glue (which can distinguish the size of 0.1~100bp) was used for electrophoresis, and the enzyme activity was verified. As can be seen from the result, the dsDNA fragment at the sticky end is mended by T4 DNA Polymerase, which has a larger molecular weight, and the electrophoretic band position is above the DNA fragment with the sticky end.
      Lane 1 Negative Control
      Lane 2 UA070070-T4 DNA Polymerase 10ng
      Lane 3 UA070070-T4 DNA Polymerase 20ng
      Lane 4 UA070070-T4 DNA Polymerase 40ng
      Lane 5 UA070070-T4 DNA Polymerase 80ng
      Lane 6 UA070070-T4 DNA Polymerase 0.1ug
      Lane 7 UA070070-T4 DNA Polymerase 0.2ug
      Lane 8 UA070070-T4 DNA Polymerase 0.4ug
      Lane 9 UA070070-T4 DNA Polymerase 0.8ug

  • 电泳

    • 2μg (R: reducing condition, N: non-reducing condition).

  • 反相高效液相色谱(RP-HPLC)

评论(0)