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. 2023 Oct 12;20:232. doi: 10.1186/s12985-023-02201-0

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 4 — Synthesis of noncanonical coronavirus transcripts is regulated under different infection conditions. A Diagram depicting the probes used for Northern blotting in Figures (B–D). B Left panel: Diagram depicting the preparation of BCoV inoculum from different passages and origins. For the BCoV inoculum from different passages, HRT-18 cells were infected with 0.1 MOI of BCoV inoculum (designated viral passage 0 BCoV; VP0 BCoV). After 48 h of infection, total cellular RNA (VP0 RNA) was harvested, and the supernatant (designated VP1 BCoV) was collected to infect another batch of fresh HRT-18 cells. For BCoV inoculum from different cell origins, HEK-293T cells were infected with 0.1 MOI of BCoV inoculum (VP0 BCoV). After 48 h of infection, the supernatant (designated VP1 HBCoV) was collected to infect another batch of HRT-18 cells at an MOI of 0.1. Right panel: Detection of noncanonical transcripts from HRT-18 cells infected with BCoV from different passages or origins by Northern blotting. The RNA species representing noncanonical transcripts between passages (Lanes 2 and 4) and origins (Lanes 4 and 6) are denoted by brown bars. C Detection of noncanonical transcripts from the same HRT-18 cells infected with different MOIs (0.1 and 10) of BCoV by Northern blotting. The RNA species representing noncanonical transcripts between different MOIs (Lanes 4 and 5) at 48 hpi are denoted by blue bars. D Comparison of the species and amounts of noncanonical transcripts between passages from fresh HRT-18 cells infected with the BCoV-p95 isolate. Left panel: Diagram depicting different passages of RNA collected from fresh HRT-18 cells infected with BCoV-p95 inoculum (VP0 BCoV-p95) and supernatant from different passages of BCoV-p95 (VP1-VP6 BCoV-p95). Right panel: Detection of noncanonical transcripts from the aforementioned total cellular RNA by Northern blotting. The RNA species representing noncanonical transcripts are denoted by orange and blue bars. kb, kilobase; m, mock-infected cells; hpi, hours postinfection; 28S, 28S rRNA