Rapid detection of environmental RNA and absolute quantification using internal strand RNA

我们已经实现了比实时PCR（qPCR）更快、更灵敏的检测，使用基于CRISPR的新检测技术，用于环境DNA和RNA快速检测的第一部分。我们分别使用针对鲤鱼设计的CRISPR和qPCR引物检测了相同环境DNA的样品，结果表明CRISPR比qPCR方法具有更高的灵敏度。此外，通过加入逆转录酶，我们可以同时检测环境DNA和RNA，将检测灵敏度提高约一个数量级。最后，我们实现了从过滤到核酸提取和最终读出的完全基于现场的一小时检测过程，该过程已被用于检测几种海洋和淡水鱼类的环境DNA和环境RNA。这些结果已经汇总并提交同行审查。我们计划通过减少反应时间，实现定量检测，简化操作步骤，使非专业人员能够使用环境DNA和RNA进行物种检测来进一步优化该方法。

We have achieved faster and more sensitive detection than real-time PCR (qPCR) using a new CRISPR-based detection technology for the first part of the rapid detection of environmental DNA and RNA. We examined samples of the same environmental DNA using the designed CRISPR and qPCR primers for Cyprinus carpio separately and showed that CRISPR possessed higher sensitivity than the qPCR method. Additionally, by adding reverse transcriptase, we can simultaneously detect environmental DNA and RNA, improving the detection sensitivity by approximately one order of magnitude. Finally, we have achieved a fully field-based, one-hour detection process from filtration to nucleic acid extraction and final readouts, which has been tested for the detection of environmental DNA and environmental RNA from several marine and freshwater fish species. These results have been summarized and submitted for peer-review. We plan to further optimize this method by reducing reaction time, achieving quantitative detection, and simplifying the operation steps to enable non-professionals to use environmental DNA and RNA for species detection.