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. 2022 Apr 23;12(5):268. doi: 10.3390/bios12050268

Figure 2.

Figure 2

SMA-Cas14a1 Fluorescence Detection System. (a) Schematic illustration of SMA-Cas14a1 fluorescence detection system. The CRISPR RNA (crRNA) targeted exon 7 of SMN1. In non-SMA individuals, SMN1 exon 7 triggers Cas14a1 activation after being recognized by the Cas14a1/crRNA complex, and activated Cas14a1 degrades the vicinal fluorescence probe nonspecifically, yielding fluorescence through acquired collateral cleavage activity. The Non-SMA individual included the SMA carrier with one copy of SMN1 exon 7 and the normal individual with more than one copy of SMN1 exon 7. However, SMA patients lack SMN1 exon 7, which is required to activate Cas14a1, and thus no fluorescence is generated in this case. (b) Schematic representation of the position and sequence targeted by crRNA1. (c) MLPA results of N-iPSCs and SMA-iPSCs. (d) Sanger sequencing results of the PCR products of N-iPSCs and SMA-iPSCs using primers F1/R1. (e) Comparison of fluorescence in different groups: 3000 ng of DNA without amplification, 1500 ng of DNA without amplification, DNA amplified using PCR, and blank control. Data are means ± SEM; **** p < 0.0001.