计算溶液所需的质量、体积或浓度。
为什么选择此级别
，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
-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。
质量文档
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。
文献证明
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。
概述
本产品是专为两步法RT-PCR第一步实验配制的cDNA第一链合成试剂盒。本产品包 含从RNA模板逆转录成cDNA第一链所需的所有试剂，其中包括HiFi-MMLV逆转录酶、 反应缓冲液、引物、dNTP等。经过突变改造的HiFi-MMLV逆转录酶RNase H活性缺失， 减少了逆转录反应中RNA的降解，更容易获得全长的cDNA。HiFi-MMLV逆转录酶热稳 定性强，可得高产量的cDNA，使用简单方便。本系统对后续的PCR以及定量PCR试 验兼容性高，适用各种PCR反应的DNA聚合酶。
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产品特点：
·RNase H-：经突变的HiFi-MMLV逆转录酶，RNase H活性缺失，更易获得全长cDNA。
·使用方便：试剂盒包含除RNA模板外的逆转录所需全部试剂。
注意事项：
1．在操作过程中应避免RNase污染，防止RNA降解或实验中的交叉污染，建议在专门 的区域进行RNA操作，使用专门的仪器和耗材，操作人员带口罩和一次性手套并经 常更换手套。
2．实验尽量使用一次性塑料器皿，若使用玻璃器皿，应使用0.1％DEPC（焦碳酸二乙 酯）水溶液在37℃处理12小时，并在120℃下高压灭菌30分钟后使用，或者将玻璃 器皿在180℃下干热灭菌60分钟后使用。实验中用到的无菌水应使用0.1%的DEPC 处理后进行高压灭菌。
3．本试剂盒中的所有试剂使用前请上下颠倒轻轻混匀，尽量避免起泡，并经短暂离心 后使用。所涉及的酶类使用后应尽快放回-20℃，避免反复冻融。
4．若起始RNA的量小于50 ng，建议加入RNA酶抑制剂（RNasin）。本试剂盒并未提供。
使用方法：
注意：10 ng-5 μg总RNA可建立20 μl反应体系，如果总RNA量大于5 μg，请按比例扩大反应体系
i 逆转录操作步骤：
1．将RNA模板、引物、dNTP Mix、DTT、RT Buffer、HiFi-MMLV和RNase-Free Water 溶解并置于冰上备用。
2．根据以下表格配制反应体系，总体积为20 μl。
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注意：
1）若起始RNA的量小于50 ng，则建议加入RNA酶抑制剂（RNasin）。本试剂盒并未提供。
2）Primer Mix由Oligo(dT)和Random Primer配制而成.
3．涡旋震荡混匀，短暂离心，使管壁上的溶液收集到管底。
4．42℃孵育30-50分钟，85℃孵育5分钟。反应结束后，短暂离心，置于冰上冷却。
5．逆转录产物可直接用于PCR反应和荧光定量PCR反应，或置于-20℃长期保存。
ii 若逆转录效率低，或RNA模板二级结构复杂、GC含量高时，建议采用以下步骤：
1．将RNA模板、引物、dNTP Mix、DTT、RT Buffer、HiFi-MMLV和RNase-Free Water 溶解并置于冰上备用。
2．根据以下表格配制反应体系，总体积为13 μl 。
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4．短暂离心，使管壁上的溶液收集到管底。
5．继续向以上反应液中加入以下试剂：
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1）若起始RNA的量小于50 ng，则建议加入RNA酶抑制剂（RNasin）。本试剂盒并未提供。
2）Primer Mix由Oligo（dT）和Random primer配制而成。
6．轻轻吹打混匀，42℃孵育50分钟，85℃孵育5分钟。
7．反应结束后，短暂离心，置于冰上冷却。
8．逆转录产物可直接用于PCR反应和荧光定量PCR反应，或置于-20℃长期保存。
This product is a cDNA first strand synthesis kit specially prepared for the first step experiment of two-step RT-PCR. This product contains all the reagents required for reverse transcription from RNA templates to cDNA first strand, including HiFi MMLV reverse transcriptase, reaction buffer, primers, dNTP, etc. The mutated HiFi MMLV reverse transcriptase RNase H activity is deficient, reducing RNA degradation in reverse transcription reactions and making it easier to obtain full-length cDNA. HiFi MMLV reverse transcriptase has strong thermal stability and can yield high yields of cDNA, making it simple and convenient to use. This system has high compatibility with subsequent PCR and quantitative PCR experiments, and is suitable for various DNA polymerase reactions.
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Product features:
·RNase H -: Mutated HiFi MMLv reverse transcriptase with reduced RNase H activity, making it easier to obtain full-length cDNA.
·Easy to use: The reagent kit contains all the reagents required for reverse transcription, except for RNA templates.
Notes:
1. During the operation process, RNase contamination should be avoided to prevent RNA degradation or cross contamination during experiments. It is recommended to perform RNA operations in specialized areas, use specialized instruments and consumables, and have operators wear masks and disposable gloves, and frequently change gloves.
2. Disposable plastic containers should be used as much as possible for experiments. If glass containers are used, they should be treated with a 0.1% DEPC (diethyl pyrocarbonate) aqueous solution at 37 ℃ for 12 hours, and sterilized under high pressure at 120 ℃ for 30 minutes before use. Alternatively, glass containers should be sterilized under dry heat at 180 ℃ for 60 minutes before use. The sterile water used in the experiment should be treated with 0.1% DEPC and then subjected to high-pressure sterilization.
3. All reagents in this reagent kit should be gently mixed upside down before use, avoiding foaming as much as possible, and used after brief centrifugation. The enzymes involved should be returned to -20 ℃ as soon as possible after use to avoid repeated freeze-thaw cycles.
If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNAsin). This kit is not provided.
Usage:
Attention: 10 ng-5 μ G Total RNA can establish 20 μ Reaction system, if the total RNA content is greater than 5 μ g. Please expand the reaction system proportionally
i Steps for reverse transcription:
1. Dissolve RNA templates, primers, dNTP Mix, DTT, RT Buffer, HiFi MMLV, and RNase Free Water and place on ice for later use.
2. Prepare a reaction system according to the following table, with a total volume of 20 μ L.
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Attention:
1) If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNAsin). This kit is not provided.
2) Primer Mix is formulated from Oligo (dT) and Random Primer
3. Vortex shake and mix well, briefly centrifuge to collect the solution on the pipe wall to the bottom of the pipe.
4.
Incubate at 42 ℃ for 30-50 minutes and 85 ℃ for 5 minutes. After the reaction is complete, centrifuge briefly and cool on ice.
5. Reverse transcripts can be directly used for PCR reactions and fluorescence quantitative PCR reactions, or stored at -20 ℃ for a long time.
ii If the reverse transcription efficiency is low, or the RNA template secondary structure is complex and the GC content is high, the following steps are recommended:
1. Dissolve RNA templates, primers, dNTP Mix, DTT, RT Buffer, HiFi MMLV, and RNase Free Water and place on ice for later use.
2. Prepare the reaction system according to the following table, with a total volume of 13 μ L.
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3.
Incubate
at 70 ℃ for 10 minutes and quickly ice bath for 2 minutes.
4. Centrifuge briefly to collect the solution on the tube wall to the bottom of the tube.
5. Continue to add the following reagents to the above reaction solution:
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Attention:
1) If the initial amount of RNA is less than 50 ng, it is recommended to add RNA enzyme inhibitors (RNAsin). This kit is not provided.
2) Primer Mix is formulated from Oligo (dT) and Random primer.
6. Gently blow and mix well, incubate at 42 ℃ for 50 minutes, and incubate at 85 ℃ for 5 minutes.
7. After the reaction is complete, centrifuge briefly and cool on ice.
8. Reverse transcripts can be directly used for PCR reactions and fluorescence quantitative PCR reactions, or stored at -20 ℃ for a long time.
规格
别名
HiFi-MMLV cDNA试剂盒
英文名称
HiFi-MMLV cDNA Kit
储存条件
-20°C储存,避免反复冻融
运输条件
超低温运输
产品线
生物产品
名称和识别符
分子类型
试剂盒
技术文档
高级数据
质检证书（CoA,COO,BSE/TSE 和分析图谱)
C of A & Other Certificates(BSE/TSE, COO):
输入批号以搜索分析图谱:
溶液计算器
常见问题
本产品应如何储存？
请于?20 °C条件下保存。避免反复冻融。建议按单次用量分装保存。
本产品如何运输？
本产品采用保温箱配冰袋运输。收货后请立即拆包，并按上述储存条件保存。
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每个订单都会随附分析证书（COA），您也可以从账户订单历史中下载。如需订购前的COA，请联系您的客户代表，或使用PDP上的"申请COA"按钮——我们将提供代表性批次的COA。
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