Extended Data Fig. 2: Testing of RBD loop-fusion variants.

Each of the fusion proteins was assembled with the crRNA as described in Methods. a–c, Testing of seven candidate RBDs (RBD#1–RBD#7) fused to LwaCas13a. a and b, Raw fluorescence over a course of 120 min in the reactions with or without 10 pM synthetic RNA targets: a, SARS-CoV-2 RdRp gene fragment (T2); b, ssRNA1 (T3). NTC, non-target control. c, The 120-min background-subtracted fluorescence from a and b. d–f, Testing of RBD#3 and RBD#4 fused to various positions on Loop 1 of LwaCas13a. The six variants were RBD#3 and #4 inserted after N415, N416, and K417, respectively. d and e, Raw fluorescence data over a course of 120 min in the reactions with or without 10 pM synthetic RNA targets: d, SARS-CoV-2 RdRp gene fragment (T2); e, ssRNA1 (T3). f, The 30-min background-subtracted fluorescence from d and e. g and h, Testing of tandem RBD insertions. The four variants for comparison were RBD#3, RBD#4, RBD#3–linker–RBD#4, and RBD#4–linker–RBD#3 inserted after N415 on Loop 1 of LwaCas13a, respectively. A 10-aa flexible linker (SGGSGGSGGS) was used to connect two RBDs in tandem. g, Background-subtracted fluorescence data over a course of 60 min in the reactions with 10 pM SARS-CoV-2 N gene fragment synthetic RNA targets (T1). h, The 30-min and 60-min background-subtracted fluorescence from g. a, b, d, e, and g, Lines and error bars represented mean ± s.e.m. from four technical replicates; c, f, and h, bar plots were expressed as mean ± s.e.m. from four technical replicates. Two-tailed p values were calculated using unpaired t-tests with Welch’s correction: ns, not significant, ^*p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001 (see SourceData for p value).

Source data