T4 DNA Polymerase | NEB酶试剂 New England Biolabs

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T4 DNA Polymerase

We are excited to announce that all reaction buffers are now BSA-free. NEB began switching our BSA-containing reaction buffers in April 2021 to buffers containing Recombinant Albumin (rAlbumin) for restriction enzymes and some DNA modifying enzymes. Find more details at www.neb.com/BSA-free.

T4 DNA Polymerase catalyzes the synthesis of DNA in the 5´→ 3´ direction and requires the presence of template and primer. 

  • Gap filling (no strand displacement activity)
  • Removal of 3’ overhangs or fill-in of 5’ overhangs to form blunt ends
  • Lacks 5’ —> 3’ exonuclease activity
  • Probe labeling using replacement synthesis
  • Singe-strand deletion subcloning

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T4 DNA Polymerase | NEB酶试剂 New England Biolabs

DNA Blunting Tutorial

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库存
货号 浓度 规格 目录价 北京 上海 广州 成都 苏州
M0203S 3,000 units/ml 150 units ¥749.00 有 有 有 有 有
M0203L 3,000 units/ml 750 units ¥2,979.00 有 有 有 无 有
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产品信息

T4 DNA Polymerase catalyzes the synthesis of DNA in the 5´→ 3´ direction and requires the presence of template and primer. This enzyme has a 3´→ 5´ exonuclease activity which is much more active than that found in DNA Polymerase I (E. coli).  Unlike E. coli DNA Polymerase I, T4 DNA Polymerase does not have a 5´→ 3´ exonuclease function.

产品来源

Purified from a strain of E. coli that carries the T4 DNA Polymerase gene.

产品类别:
DNA Manipulation Products

应用:
Blunting,
Polymerases for DNA Manipulation,
PCR

  • 产品组分信息

    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
    • M0203S     -20    
        T4 DNA Polymerase M0203SVIAL -20 1 x 0.05 ml 3,000 units/ml
        NEBuffer™ r2.1 B6002SVIAL -20 1 x 1.25 ml 10 X
    • M0203L     -20    
        T4 DNA Polymerase M0203LVIAL -20 1 x 0.25 ml 3,000 units/ml
        NEBuffer™ r2.1 B6002SVIAL -20 1 x 1.25 ml 10 X

  • 特性和用法

    单位定义

    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 (5).

    反应条件

    1X NEBuffer™ r2.1

    1X NEBuffer™ r2.1
    50 mM NaCl
    10 mM Tris-HCl
    10 mM MgCl2
    100 µg/ml Recombinant Albumin
    (pH 7.9 @ 25°C)

    在不同缓冲液中的活性

    NEBuffer™ 1: 60%
    NEBuffer™ 2: 100%
    NEBuffer™ 3: 100%
    NEBuffer™ 4: 100%

    贮存溶液

    100 mM KPO4
    1 mM DTT
    50% Glycerol
    pH 6.5 @ 25°C

    热失活

    75°C for 20 min

    分子量

    理论上的: 104000 daltons

    5′ – 3′ 核酸外切酶

    No

    3′ – 5′ 核酸外切酶

    Yes

    链置换

    No

    单位活性检测条件

    1X NEBuffer 2.1, 33 µM dNTPs including [3H]-dTTP, 70 µg/ml denatured herring sperm DNA.

    错配率

    ~ 1×10-6bases

  • 优势和特性

    应用特性

    • 3´ overhang removal to form blunt ends (1,2).
    • 5´ overhang fill-in to form blunt ends (1,2).
    • Single strand deletion subcloning (3).
    • Second strand synthesis in site-directed mutagenesis (4).
    • Probe labeling using replacement synthesis (1,2).

  • 相关产品

    相关产品

    • Deoxynucleotide (dNTP) Solution Set
    • Deoxynucleotide (dNTP) Solution Mix
    • BSA，分子生物学级
    • Monarch® Plasmid Miniprep Kit 
    • Monarch® DNA Gel Extraction Kit
    • Monarch® PCR & DNA Cleanup Kit (5 μg)

    单独销售的组分

    • NEBuffer™ r2.1

  • 注意事项
    1. For fill-in reactions only: T4 DNA Polymerase can be used in NEBuffers 1.1, 2.1, 3.1 and CutSmart® Buffer as well as NEBuffers 1-4 and T4 DNA Ligase Reaction Buffer.
    2. For blunting reactions requiring removal of overhangs: T4 DNA Polymerase can be used in NEBuffers 1.1, 2.1, and CutSmart Buffer as well as NEBuffers 1, 2, and 4 and T4 DNA Ligase Reaction Buffer. NEBuffers 3.1 and 3 are not recommended when overhang removal is required.
    3. Optimal activity is observed in NEBuffer 2.1.
    4. Supplement with dNTPs.*
    5. BSA supplementation is recommended when using a buffer that does not already contain BSA.
    6. Incubate at temperature suggested for specific protocol.

      * Refer to specific protocol to determine recommended dNTP concentrations.

      Refer

  • 参考文献
    1. Tabor, S. and Struhl, K. (1989). DNA-Dependent DNA Polymerases. In F. M. Ausebel, R. Brent, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith and K. Struhl(Ed.), Current Protocols in Molecular Biology. 3.5.10-3.5.12. New York: John Wiley & Sons, Inc.
    2. Sambrook, J. et al. (1989). Molecular Cloning: A Laboratory Manual. (2nd ed.), 5.44-5.47. Cold Spring Harbor: Cold Spring Harbor Laboratory Press.
    3. Dale, R. et al. (1985). Plasmid. 13, 31-40.
    4. Kunkel, T.A. et al. (1987). Methods Enzymol.. 154, 367-382.
    5. Panet, A. et al. (1973). Biochemistry. 12, 5045-5050.

操作说明、说明书 & 用法

  • 操作说明
    1. Protocol for blunting ends by 3’ overhang removal and 3’ recessed (5’ overhang) end fill-in using T4 DNA Polymerase (M0203)

  • 使用指南
    • Activity of DNA Modifying Enzymes in rCutSmart™ Buffer

工具 & 资源

  • 选择指南
    • Blunting Selection Chart
    • Common Applications for Exonucleases and Endonucleases
    • DNA Polymerase Selection Chart

  • Web 工具
    • NEBcloner®

FAQs & 问题解决指南

  • FAQs
    1. Can T4 DNA Polymerase be used in other NEBuffers and rCutSmart Buffer?
    2. Can T4 DNA Polymerase be used to blunt DNA?
    3. Can T4 DNA Polymerase be used to fill in 3′ overhangs?
    4. Can T4 DNA Polymerase be used to remove 5′ overhangs?
    5. Can T4 DNA Polymerase be heat inactivated?
    6. Are the nucleotides needed to remove a 3′ overhang using T4 DNA Polymerase?
    7. What are the main causes of blunting reaction failure using T4 DNA Polymerase?
    8. Can T4 DNA Polymerase be used in labeling reactions and partial fill in reactions?
    9. Is T4 DNA Polymerase active at room temperature?
    10. Is T4 DNA Polymerase the enzyme of choice for removing 3′ overhangs and filling in 5′ overhangs (3′ recessed ends)?
    11. Are NEB DNA Polymerases supplied with dNTPs?
    12. Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?

  • 实验技巧
    Excess enzyme, temperatures above 12°C, or limited dNTP concentrations can cause excessive 3’ → 5’ exonuclease degradation, which eliminates blunt end formation.

引用 & 技术文献

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质控、安全 & 法规

  • 质控分析
    每一新批次的 NEB 产品都要经过质控，以满足指定的质量标准，对特定产品的质量标准和每一批次的检测数据可以通过产品说明表格、COA、产品信息卡或者产品手册进行查阅或下载。关于 NEB 产品质控的详细信息，可从此处查阅。

  • 产品说明与变更通知

    产品说明与变更通知

    产品说明表包含产品的储存温度、有效期和规格，这些文件的命名规则如下：[货号]_[规格]_[版本]

    • M0203S_L_v1

  • CoA
    CoA 文件包含单个批次产品的储存温度、有效期和质控，这些文件的命名规则如下： [货号]_[规格]_[版本]_[Lot#]

    • M0203S_L_v1_0401512
    • M0203S_L_v1_0401606
    • M0203S_L_v1_0401612
    • M0203S_L_v1_0401706
    • M0203S_L_v1_0401712
    • M0203S_v1_10011307
    • M0203S_v1_10021832
    • M0203L_v1_10019934
    • M0203L_v1_10023074
    • M0203S_v1_10029314
    • M0203L_v1_10034499
    • M0203S_v1_10034498
    • M0203L_v1_10036401
    • M0203L_v1_10043962
    • M0203S_v1_10043963
    • M0203L_v1_10048109
    • M0203S_v1_10049314
    • M0203S_v1_10054179
    • M0203L_v1_10055853
    • M0203S_v1_10055855
    • M0203L_v1_10060120
    • M0203S_v1_10060122
    • M0203L_v1_10063501
    • M0203S_v1_10063553
    • M0203L_v1_10068639
    • M0203S_v1_10070264
    • M0203L_v1_10074951
    • M0203S_v1_10075027
    • M0203S_v1_10081947
    • M0203S_v1_10085012
    • M0203L_v1_10085011
    • M0203S_v1_10091124
    • M0203S_v1_10091904
    • M0203L_v1_10091907
    • M0203L_v1_10099938
    • M0203S_v1_10099937
    • M0203L_v1_10107643
    • M0203S_v1_10107644
    • M0203S_v1_10112939
    • M0203L_v1_10113492
    • M0203L_v1_10127968
    • M0203S_v1_10127969
    • M0203S_v1_10143141
    • M0203L_v1_10145960
    • M0203L_v1_10151704
    • M0203S_v1_10151890

  • SDS
    以下 SDS 文件可以帮助您安全地使用该产品

    • T4 DNA Polymerase
    • NEBuffer™ r2.1