CRISPR-Cas12b-assisted nucleic acid detection platform

Abstract Rapid molecular diagnostic technology is very useful in many areas, including public health, environmental testing and criminal investigation. We recently showed that Cas12a had trans -cleavage activity upon collateral single-stranded DNA (ssDNA), with which the HOLMES platform (one- HO ur L ow-cost M ultipurpose highly E fficient S ystem) was developed. Here, we combine the thermophilic Cas12b, which also has the ssDNA trans -cleavage activity, with Loop-Mediated Isothermal Amplification (LAMP), and create HOLMESv2. In HOLMESv2, LAMP amplification and Cas12b trans -cleavage can be integrated into a one-step system with a constant temperature, which therefore brings much convenience in nucleic acid detection. Moreover, we also simplify the RNA detection procedures in HOLMESv2, using an RNA-dependent DNA polymerase for amplification and therefore omitting an extra reverse transcription step. One Sentence Summary We combine LAMP and Cas12b to develop HOLMESv2 for conveniently detecting target nucleic acid in a one-step approach.