Vol.:(0123456789) 1 3 Theoretical and Applied Genetics https://doi.org/10.1007/s00122-019-03341-0 ORIGINAL ARTICLE CRISPR/Cas9‑mediated genome editing reveals diferences in the contribution of INDEHISCENT homologues to pod shatter resistance in Brassica napus L. Yungu Zhai 1  · Shengli Cai 1  · Limin Hu 1  · Yang Yang 1  · Olalekan Amoo 1  · Chuchuan Fan 1  · Yongming Zhou 1 Received: 31 December 2018 / Accepted: 5 April 2019 © Springer-Verlag GmbH Germany, part of Springer Nature 2019 Abstract The INDEHISCENT (IND) and ALCATRAZ (ALC) gene homologues have been reported to be essential for dehiscence of fruits in Brassica species. But their functions for pod shatter resistance in Brassica napus, an important oil crops, are not well understood. Here, we assessed the functions of these two genes in rapeseed using CRISPR/Cas9 technology. The induced mutations were stably transmitted to successive generations, and a variety of homozygous mutants with loss-of-function alleles of the target genes were obtained for phenotyping. The results showed that the function of BnIND gene is essential for pod shatter and highly conserved in Brassica species, whereas the BnALC gene appears to have limited potential for rapeseed shatter resistance. The homoeologous copies of the BnIND gene have partially redundant roles in rapeseed pod shatter, with BnA03.IND exhibiting higher contributions than BnC03.IND. Analysis of data obtained from the gene expres- sion and sequence variations of gene copies revealed that cis-regulatory divergences alter gene expression and underlie the functional diferentiation of BnIND homologues. Collectively, our results generate valuable resources for rapeseed breeding programs, and more importantly provide a strategy to improve polyploid crops. Introduction Rapeseed (Brassica napus L., AACC, 2n = 38) is the sec- ond largest oil crop after soybean globally (https://www.ers. usda.gov/topics/crops/soybeans-oil-crops/canola), account- ing for about 16% of the total global vegetable oil production (Hu et al. 2017; Woodfeld et al. 2017). It provides edible oil for human diets, protein-rich feed for animals and raw materials for industrial processes, such as biodiesel produc- tion. Achieving high yields and genetic improvements have always been the major goals in rapeseed production. How- ever, pod shatter often occurs after rapeseed maturation and it is one of the major bottlenecks in rapeseed production worldwide. The resulting loss in yield usually accounts for about 20% of total production and up to 50% under adverse weather conditions (Price et al. 1996; Child et al. 1998). Moreover, shattered seeds can remain viable in soil for sev- eral years and become volunteer plants in subsequent crops in the rotation cycle, increasing weed control problems (Morgan et al. 2000). Therefore, rapeseed is often harvested ahead of maturity to avoid seed loss. However, this results in chlorophyll-contaminated oil extracted from immature seed and lowering its quality (Hu et al. 2015). Pod shatter is also a critical factor afecting mechanized harvesting in rapeseed, Communicated by Maria Laura Federico. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00122-019-03341-0) contains supplementary material, which is available to authorized users. * Chuchuan Fan fanchuchuan@mail.hzau.edu.cn Yungu Zhai zhaiyungu@webmail.hzau.edu.cn Shengli Cai tim@webmail.hzau.edu.cn Limin Hu hulimin@webmail.hzau.edu.cn Yang Yang yangyangyy@webmail.hzau.edu.cn Olalekan Amoo olalekan@webmail.hzau.edu.cn Yongming Zhou ymzhou@mail.hzau.edu.cn 1 National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China