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. 2018 Nov 21;8:17195. doi: 10.1038/s41598-018-35479-0

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© The Author(s) 2018

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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Two-Step ORNi-PCR using cDNA to discriminate splice variants. (A) Schematic diagram of Pax5-1B cDNA prepared from DT40 cells. Two forms of transcripts including exon 1B are produced from the chicken Pax5 gene in DT40 cells. A part of the exon 1B region is truncated in the shorter form (truncated). (B) Target position of ORN_cPax5_Ex1B, a 20 base ORN targeting exon 1B of the chicken Pax5 locus. The cDNA sequence of Pax5-1B (full-length) is shown. The underlined sequence is truncated in Pax5-1B (truncated) cDNA. (C) Conditions for Two-Step ORNi-PCR using cDNA. (D) Results of Two-Step ORNi-PCR. The predicted positions of each amplicon are indicated by arrows (full-length and truncated forms). M, molecular weight markers.