TWISTDX RPA试剂盒（（加MMLV和抑制剂）操作建议

英国TwistDx公司（前身为ASM科技有限公司）成立于1999年，基于英国剑桥Babraham研究院建立的生物技术公司。该公司通过突破等温DNA扩增，开发的重组酶聚合酶扩增*技术（RPA），被誉为DNA诊断领域的革命性创新。

RT试剂盒举例

（1）Revert Reverse Transcriptase(200U/uL)(Thermo EP0441)

（2）RNA Inhibitor (40,000 U/mL)( Enzymatics Y9240L

TWISTDX厂家回复

　　“We recommend using similar RT amounts to that of a PCR reaction of the same volume, according to your chosen manufacturer’s instructions.

　　We also suggest that you run reactions at 40°C and delay agitation of the reaction by a minute to give the RT time to work, otherwise you just run the reaction as you would a normal TwistAmp® reaction (it’s a one-step process).

　　It’s also advisable to add RNase Inhibitor to any reaction where RNA is the target material. Any commercially available RNase Inhibitor will be suitable.

　　If the M-MLV RT that you are using comes with a separate dilution/reaction buffer, we suggest not using it as it will lower the concentration of the enzyme.

　　The “delay agitation of the reaction by a minute” refers to the first mixing step per the protocol in the manual (Step 4). So wait 5 minutes instead of 4 minutes before “vortex, spin down briefly, and return the samples to the reader ensuring that the tubes are returned to their original positions in the block.”

　　If we were to try and use it again, we would do a titration to determine optimal quantity to add in the new formulation reactions (we haven’t used it since we changed the formulation of the reactions a couple of years ago).

　　When using RNase Inhibitor. We have used 1µg /50µl reaction, some R&D users have used 2µg /50ul, which didn’t appear to have a negative effect on the performance in our own assay.

　　For this : RNAse Inhibitor, that manufacture uses 10 ul of the RNAse solution in 50 ul reaction volumes for activity determination, but your customer can try different amounts 5ul or 20ul to see what will give the best results. For this product, references of 50 ul PCR reaction volumes would also be reasonable.

TwistDx以此技术为基础生产的核酸扩增类产品，能在恒定且较低的温度下，仅用10~20分钟，进行单分子核酸检测。这一技术对环境、硬件设施的要求很低，对于生物防护、水体检验、食品检验、医疗诊断、微流体、兽医等方面都有良好的应用。

RPA扩增的各种体系试剂盒中，含有DNA、RNA扩增所需的所有试剂，研究者只要准备好引物和模板就行了。扩增结果可以分别通过凝胶电泳检测、荧光定量检测或试纸条检测，产物纯化后可用于下游研究。

默瑞生物

默瑞(上海)生物科技有限公司，作为专业的生命科学产品推广与服务商，以“创新，专业，共赢”的理念，不断强化服务体系，提升客户体验，以莫纳生物产品为主线，结合国内外优质产品提供高标准完整解决方案，帮助科研客户和生物技术企业高效创新，加速转化，创造更多优质产品，提升社会福祉，实现多方共赢。

　　默瑞生物创立以来，已经陆续导入TwistDx,enzymatics,KAPABIOSYSTEMS,BioDynami,Jackson,Echelon,ChromoTek,Nanocs,Biotechrabbit,LeeBiosolutions, magtivio等优质品牌，逐步培育成为专业用户的选择。