Single-nucleotide polymorphisms (SNPs) are of crucial importance in genetics, disease association studies, and personalized medicine. However, traditional detection methods (such as sequencing and PCR) are time-consuming and have limited single-base resolution. Recently, the research group led by Associate Professor Liu Yizhen from Shenzhen University reported a novel SNP genotyping technology, which enables rapid single-tube isothermal detection by integrating recombinase polymerase amplification (RPA), Lambda exonuclease, and the CRISPR-Cas13a system. Recently, the related achievements were published in Biosensors and Bioelectronics, an authoritative international journal in the field of biosensing, under the title "Single-tube Lambda exonuclease-mediated LbuCas13a detect of ssDNA for single-nucleotide polymorphisms genotyping" (DOI: https://doi.org/10.1016/j.bios.2025. 117760). Luo Siyuan, a 2022 master's student majoring in Chemistry, Associate Researcher Chen Yong, and Li Zongnan, a 2022 undergraduate student majoring in Chemistry, are the first authors. Associate Professor Liu Yizhen is the corresponding author, and Shenzhen University is the sole affiliated institution.

In this study, a single-tube isothermal SNP genotyping technology named SNP-SENSE was developed. By integrating recombinase polymerase amplification (RPA), Lambda exonuclease, and the CRISPR-LbuCas13a system, it enables rapid and highly specific detection of single-nucleotide polymorphisms within 30 minutes.

Detection results from actual whole blood samples confirmed that the CYP2C19 *2/*3/*17 genotyping results of 20 samples using SNP-SENSE were 100% consistent with those obtained by sequencing. 

The SNP-SENSE platform established in this study achieves, for the first time, RPA amplification, Lambda exonuclease digestion, and LbuCas13a detection in a single tube, completing SNP genotyping within 30 minutes under a constant temperature of 39°C. Its core innovation lies in utilizing the ultra-high single-base resolution of LbuCas13a for ssDNA, combined with the specific cleavage of Lambda exonuclease to generate ssDNA, eliminating the need for transcription steps or large-scale temperature-controlled instruments. The SNP-SENSE platform demonstrated aM-level sensitivity and 100% accuracy in both simulated samples and clinical blood tests, addressing the issues of time-consuming procedures, susceptibility to contamination, and insufficient single-base discrimination of traditional methods. It provides an efficient tool for point-of-care genetic testing and personalized medicine. 

This research was supported by the Shenzhen Special Fund for Medical Research, the National Natural Science Foundation of China, the Guangdong Basic and Applied Basic Research Foundation, and the Shenzhen Natural Science Foundation.