Vol.:(0123456789) 1 3 Theoretical and Applied Genetics https://doi.org/10.1007/s00122-019-03396-z ORIGINAL ARTICLE Next‑generation sequencing‑based bulked segregant analysis for QTL mapping in the heterozygous species Brassica rapa Noriaki Itoh 1  · Tenta Segawa 1  · Muluneh Tamiru 2  · Akira Abe 3  · Shota Sakamoto 1  · Aiko Uemura 3  · Kaori Oikawa 3  · Hiroto Kutsuzawa 1  · Hironori Koga 1  · Tomohiro Imamura 1  · Ryohei Terauchi 3,4  · Hiroki Takagi 1 Received: 27 February 2019 / Accepted: 6 July 2019 © Springer-Verlag GmbH Germany, part of Springer Nature 2019 Abstract Key message An improved protocol of QTL-seq, an NGS-based method for bulked segregant analysis we previously developed in rice, allowed successful mapping of QTLs of interest in the highly heterozygous genome of B. rapa, demonstrating the power of this elegant method for genetic analyses in heterozygous species of economic importance. Abstract Recent advances in next-generation sequencing (NGS) and the various NGS-based methods developed for rapidly identifying candidate genes of interest have accelerated genetic analysis mainly in the model plants rice and Arabidopsis. Brassica rapa includes several economically important crops such as Chinese cabbage, turnip and various leafy vegetables. The application of NGS-based approaches for the analysis of B. rapa has been limited mainly due to its highly heterozy- gous genome and poor quality of the reference genome sequence currently available for this species. In this study, we have improved QTL-seq, a method for NGS-based bulked segregant analysis we previously developed in rice, extending its applicability for accelerating the genetic analysis and molecular breeding of B. rapa. Addition of new flters to the original QTL-seq pipeline allowed removal of spurious single-nucleotide polymorphisms caused by alignment/sequencing errors and variability between parents, signifcantly improving accuracy of the analysis. As proof of principle, we successfully applied the new approach to identify candidate genomic regions controlling fowering and trichome formation using segregating F2 progeny obtained from crosses made between cultivars of B. rapa showing contrasting phenotypes for these traits. We strongly believe that the improved QTL-seq method reported here will extend the applicability of NGS-based genetic analysis not only to B. rapa but also to other plant species of economic importance with heterozygous genomes. Introduction Brassica rapa is independently domesticated in Europe and Asia, from where its cultivation spread worldwide (Zhao et al. 2005; Cheng et al. 2016). Its long history of cultiva- tion in diferent regions of the world has led to considerable genetic diversity because of both natural selection (adapta- tion to variable environments) and selective breeding that targeted manipulating size of harvestable plant parts to meet consumer preferences in diverse cultures (Yarkhunova et al. 2016). B. rapa represents many economically important crops such as Chinese cabbage (ssp. pekinensis), Pak choi (ssp. chinensis), Wutacai (ssp. narinosa), oil seed types (ssp. oleifera) and turnip (ssp. rapa) that have been subjected to diferent breeding strategies (Diederichsen 2001). However, B. rapa breeding is yet to fully beneft from the various NGS-based techniques that have accelerated crop improve- ment in multiple crop species (Zhao 2015). Communicated by Albrecht E. Melchinger. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00122-019-03396-z) contains supplementary material, which is available to authorized users. * Hiroki Takagi h-takagi@ishikawa-pu.ac.jp 1 Ishikawa Prefectural University, 1-308, Suematsu, Nonoichi, Ishikawa 921-8836, Japan 2 Centre for AgriBioscience (AgriBio), La Trobe University, 5 Ring Road, Bundoora, VIC 3086, Australia 3 Iwate Biotechnology Research Center, 22-174-4, Narita, Kitakami, Iwate 024-0003, Japan 4 Kyoto University, Nakajou 1, Mozume, Mukou, Kyoto 617-0001, Japan