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. 2022 Aug 10;13:4691. doi: 10.1038/s41467-022-32269-1

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© The Author(s) 2022

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a Schematic representation of the CA_C0764 gene with a group II intron inserted at position 408 on the sense strand of CA_C0764. b PCR screening for the identification of putative CA_C0764-408s::CT mutants using gene-specific primers flanking the intron insertion site (lanes 2–5) (expected size 2060 bp) and PCR control with wild-type DNA (lane 7) (expected size 270 bp), lanes 1 and 8 DNA ladder. c Southern hybridization to demonstrate the presence of a single intron insertion in the selected MGCΔcac1502cac0764-408s::CT mutant. The intron probe was DIG-labeled and hybridized to HindIII-HF digested genomic DNA of the MGCΔcac1502cac0764-408s::CT mutant (lane 2) with an expected size of 1970 bp. The HindIII-HF digested genomic DNA of the C. acetobutylicum MGCΔcac1502 strain (lane 1) was also tested as a negative control. Lane 3 is a 1 kb DNA ladder. d PCR screening for the identification of one putative gltb181s::CT mutant (among three colonies that were tested) using gene-specific primers flanking the intron insertion site (lanes 1–3) (expected size 2200 bp) and PCR control with wild-type DNA (lane 4) (expected size 500 bp), lane 1 DNA ladder. Source data are provided as a Source Data file.