EnzymoPure™ 级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
-20°C储存,避免反复冻融。超低温运输 。请查阅批次 COA 获取详细规格。
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
mTaq DNA Polymerase是通过缺失Taq DNA Polymerase N端一段氨基酸和突变 改造得到的新型DNA聚合酶。通过改造后使本产品能够全血中存在的抑制剂产生耐受 作用，能够直接扩增人和小鼠全血样品中的DNA而无需事先对基因组进行提取和纯化。 PCR产物3’端为A，可直接用于T/A克隆。
Notice: mTaq PCR Buffer contains 30 mM MgCl₂ |
质量控制：
经过多次柱纯化，SDS-PAGE检测其纯度大于99%；经检测无外源核酸酶活性； PCR方法检测无宿主残余DNA；能有效地扩增人基因组中的单拷贝基因；室温存放一 周，无明显活性改变。
使用方法：
1. 使用前请将mTaq DNA Polymerase反复颠倒至完全混匀。
2. 将PCR薄壁管置于冰上，加入除全血外的以下试剂。
|
注意：
1）*加入全血前反复上下吸打完全混匀各种试剂、
2）DNA模板：可以使用肝素钠、Na-EDTA、K-EDTA或柠檬酸钠处理全血。通常建议全血含量 为5-10%。不推荐使用高浓度血液。对于高GC含量的模板，加入10%DMSO。
3）引物：寡核苷酸引物长度通常含20-30个核苷酸，并且最好GC含量在40-60%并均匀分布于引 物中。在常规的PCR反应中，引物浓度请以终浓度0.1-1.0 μM作为设定范围的参考。
3. 最后将全血加入管底。
4. PCR反应条件
|
注意：
1）PCR仪于94-95℃预热，将样品放置于PCR仪上开始循环。
2）mTaq提高了冷敏感性，具备了一些热启动特性。通常可以通过在冰上配制反应成分、最后 加入聚合酶以及将热循环仪预热至变性温度（95℃）后立即进行反应来避免非特异性产物的 产生。
3）变性温度与时间：在PCR循环前为了充分裂解血液细胞和释放/变性DNA，要求初始变性为 95℃ 5分钟。
4）退火温度与时间:退火时间通常为30秒-1分钟。退火温度可以低于理论退火温度（Tm）5℃开 始，通过梯度PCR进行优化。
5）延伸时间：延伸反应通常在72℃下进行。一般每250-500 bp延伸时间为1分钟。推荐72℃ 10 分钟进行最终延伸。
6）通常35-40个循环可以达到最优扩增。
5. 结果检测：反应结束后取5 µl反应产物并加上电泳缓冲液一起电泳检测结果。
MTaq DNA Polymerase is a novel DNA polymerase obtained by deletion of a segment of amino acid at the N-terminus of Taq DNA Polymerase and mutation modification. Through modification, this product can produce tolerance to inhibitors present in whole blood, allowing for direct amplification of DNA in human and mouse whole blood samples without the need for prior genome extraction and purification. The 3 'end of the PCR product is A and can be directly used for T/A cloning.
Notice: mTaq PCR Buffer contains 30 mM MgCl₂ |
Quality control:
After multiple column purifications, SDS-PAGE detected a purity of over 99%; No exogenous nuclease activity detected; PCR method for detecting residual DNA without host; Can effectively amplify single copy genes in the human genome; Store at room temperature for one week without significant changes in activity.
Usage:
1. Before use, please invert the mTaq DNA Polymerase repeatedly until completely mixed.
2. Place the PCR thin-walled tube on ice and add the following reagents except for whole blood.
|
Attention:
1) Before adding whole blood, repeatedly aspirate and beat up and down to completely mix various reagents
2) DNA template: Whole blood can be treated with sodium heparin, Na EDTA, K-EDTA, or sodium citrate. It is usually recommended to have a whole blood content of 5-10%. It is not recommended to use high concentration blood. For templates with high GC content, add 10% DMSO.
3) Primers: Oligonucleotide primers typically contain 20-30 nucleotides in length, with a GC content of 40-60% and evenly distributed within the primers. In conventional PCR reactions, the primer concentration should be at the final concentration of 0.1-1.0 μ M serves as a reference for setting the range.
3. Finally, add whole blood to the bottom of the tube.
4. PCR reaction conditions
|
Attention:
1) Preheat the PCR instrument at 94-95 ℃ and place the sample on the PCR instrument to start cycling.
2) MTaq enhances cold sensitivity and has some hot start characteristics. Usually, the production of non-specific products can be avoided by preparing reaction ingredients on ice, adding polymerase, and preheating the thermal cycler to denaturation temperature (95 ℃) for immediate reaction.
3) Denaturation temperature and time: In order to fully lyse blood cells and release/denature DNA before the PCR cycle, an initial denaturation temperature of 95 ℃ for 5 minutes is required.
4) Annealing temperature and time: The annealing time is usually 30 seconds to 1 minute. The annealing temperature can be lower than the theoretical annealing temperature (Tm) by starting at 5 ℃ and optimizing through gradient PCR.
5) Extension time: The extension reaction is usually carried out at 72 ℃. Generally, the extension time is 1 minute every 250-500 bp. Recommend final extension at 72 ℃ for 10 minutes.
6) Typically, 35-40 cycles can achieve optimal amplification.
5. Result detection: After the reaction is completed, take 5 µ l of the reaction product and add it to the electrophoresis buffer for electrophoresis detection.
危险品化学品经营许可证（带存储）