32The exon junction complex (EJC) is an essential constituent and regulator of spliced messenger 33 ribonucleoprotein particles (mRNPs) in metazoans. As a core component of the EJC, CASC3 was 34 described to be pivotal for EJC-dependent nuclear and cytoplasmic processes. However, recent 35 evidence suggests that CASC3 functions differently from other EJC core proteins. Here, we have 36 established human CASC3 knockout cell lines to elucidate the cellular role of CASC3. In the knockout 37 cells, overall EJC composition and EJC-dependent splicing are unchanged, whereas mRNA isoforms 38 targeted by nonsense-mediated decay (NMD) are upregulated on a transcriptome-wide scale. 39Mechanistically, recruiting CASC3 to reporter mRNAs by direct tethering or via binding to the EJC 40 stimulates mRNA decay and endonucleolytic cleavage at the termination codon. Building on existing 41 EJC-NMD models, we propose that CASC3 equips the EJC with the ability to communicate with the 42
The global spread of SARS-CoV-2 is a serious public health issue. Large-scale surveillance screenings are crucial but can exceed test capacities. We (A) optimized test conditions and (B) implemented pool testing of respiratory swabs into SARS-CoV-2 diagnostics. Study design: (A) We determined the optimal pooling strategy and pool size. In addition, we measured the impact of vortexing prior to sample processing, compared a pipette-pooling method (by combining transport medium of several specimens) and a swab-pooling method (by combining several swabs into a test tube filled with PBS) as well as determined the sensitivities of three PCR assays. (B) Finally, we applied high-throughput pool testing for diagnostics. Results: (A) In a low prevalence setting, we defined a preferable pool size of ten in a two-stage hierarchical pool testing strategy. Vortexing of swabs (n = 33) increased cellular yield by a factor of 2.34. By comparing Ct-values of 16 pools generated with two different pooling strategies, pipette-pooling was more efficient compared to swab-pooling. Measuring dilution series of 20 SARS-CoV-2 positive samples in three PCR assays simultaneously revealed detection rates of 85% (assay I), 50% (assay II), and 95% (assay III) at a 1:100 dilution. (B) We systematically pooled 55,690 samples in a period of 44 weeks resulting in a reduction of 47,369 PCR reactions. Conclusions: For implementing pooling strategies into high-throughput diagnostics, we recommend utilizing a pipette-pooling method, performing sensitivity validation of the PCR assays used, and vortexing swabs prior to analyses. Pool testing for SARS-CoV-2 detection is feasible and effective in a low prevalence setting.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.