Journal: Advanced Science

Article Title: CRISPR‐Scan: Cas13a Mismatch Tolerance Enabled Ultra‐Multiplexed Detection of Same‐Site Indels in Acute Myeloid Leukemia

doi: 10.1002/advs.78229

Figure Lengend Snippet: The reaction principle and workflow of the CRISPR‐Scan system. (a) Workflow of the CRISPR‐Scan detection system. Genomic DNA was extracted from patient blood samples, and 1 µL of genomic DNA was directly added to the CRISPR‐Scan reaction mixture, which contained T7 RNA polymerase, RPA reagents, a pair of primers, RNA reporter, LwaCas13a, and a single crRNA. The one‐pot reaction was performed at 39°C for 30 min inside the homemade microfluidic device. Real‐time fluorescence curves and visual detection results were recorded with a smartphone, and detection results were output automatically. (b) For CRISPR‐Scan detection, the mutant or wild‐type (WT) template was amplified using primers containing a T7 promoter sequence by RPA, followed by T7 transcription to produce RNA products. LwaCas13a/crRNA recognized RNA targets with 1–4 bp mismatches, activated its trans ‐cleavage activity, and induced cleavage of the RNA reporter, which produces fluorescence signals. However, the WT RNA contains a large number of base mismatches, making it difficult for LwaCas13a/crRNA to recognize, and therefore no fluorescence signals were generated. LwaCas13a/crRNA can achieve the distinction between WT and NPM1 gene c.863_864 4‐bp insertions, while also covering the detection of 4‐bp random insertion mutations. Created within BioRender.com. Bai, L. (2026) https://BioRender.com/12nctes .

Article Snippet: Recombinant RNase Inhibitor (Cat. No. 2313A) and Taq DNA Polymerase (Cat. No. R007A) were purchased from Takara Bio (Dalian, China).

Techniques: CRISPR, Blood/Plasma/Serum Samples, Fluorescence, Mutagenesis, Amplification, Sequencing, Activity Assay, Generated