Additional file 1 of Application of the amplification-free SERS-based CRISPR/Cas12a platform in the identification of SARS-CoV-2 from clinical samples
Creators: name: Liang, Jiajie
name: Teng, Peijun
name: Xiao, Wei
name: He, Guanbo
name: Song, Qifang
name: Zhang, Ying
name: Peng, Bin
name: Li, Gan
name: Hu, Liangshan
name: Cao, Donglin
name: Tang, Yong
Publisher: name: figshare
Year: 2021
Subjects: subject: Biochemistry
subject: Microbiology
subject: FOS: Biological sciences
subject: Cell Biology
subject: Genetics
subject: Molecular Biology
subject: Immunology
subject: FOS: Clinical medicine
subject: Biological Sciences not elsewhere classified
subject: Information Systems not elsewhere classified
subject: Cancer
subject: Mental Health
subject: Computational Biology

description: Additional file 1: Figure S1. Verification of the collateral cleavage activity of Cas12a using fluorescent assay. (a) Schematic of the Cas12a-based fluorescent assay; (b) The collateral cleavage of the Cas12a-crRNA duplex activated by dsDNA triggers. Randomly designed TargetDNAs activate respectiveCas12a-crRNA duplex. Figure S2. Confirmation of the Target DNA homologous crRNA using fluorescent assay and the optimized conditions. (a) Cas12a crRNA 1 was programmed to specifically Target DNA 1, and its time-course detection. The numbers represent the fluorescence ratio of adjacent points-in-time;(b) Cas12a crRNA 2 was programmed to specifically Target DNA 2;(c) Cas12a crRNA 3 was programmed to specifically Target DNA 3;(d) The concentrations of crRNAs and Cas12a were optimized. All the error bars are determined from three independent experiments. Figure S3. Confirmation of SARS-CoV-2 homologous crRNA. (a) Genome map of the SARS-CoV-2 showing crRNA. Visualization of the crRNA to identify N gene region in the SARS-CoV-2 genome; (b) crRNA specificity. Cas12a crRNA is programmed to specifically target SARS-CoV-2. The N gene crRNA used in the assay was specific for SARS-CoV-2 and failed to detect SARS-CoV and bat SARS-like coronavirus; (c) Time-course detection of the plasmids (2 nM) containing the N gene sequence of SARS-CoV-2, SARS-CoV and bat SARS-like coronavirus. The numbers represent the fluorescence ratio of adjacent points-in-time. All the error bars are determined from three independent experiments. Figure S4. Confirmation of the S-CRISPR assay. (a) SEM images (ZEISS ULTRA 55 field emission scanning electron microscopy) of AgNPs (І; scale bar: 200 nm; 30 K ×), SERS probe (MBs-ssDNA-AgNPs; ІІ; scale bar: 500 nm; 20 K ×), and SERS probe cleaved by Cas12a (ІІІ; scale bar: 500 nm; 20 K ×). AgNPs were successfully linked on the surfaces of the MBs and unlinked from them after cleavage; (b) The concentrations of SH-ssDNA-biotin and AgNPs@4ATP are optimized; (c) S-CRISPR assay was confirmed using Target DNA 1. All the error bars are determined from three independent experiments. Figure S5. Portable Raman plate reader. (a) The portable Raman spectrometer (QSPEC, SmartRaman); (b) Manualoperation procedureof the portable Raman spectrometer; (c) DNase-cleaving SERS probe and signals as detected by the portable Raman plate reader. All the error bars are determined from ten independent experiments. Table S1. Characteristics of clinical samples from COVID-19-suspected patients. Table S2. Nucleic acids used in this study.