- 著者
- Masahiro Nishihara Akiko Hirabuchi Fumina Goto Aiko Watanabe Chiharu Yoshida Rie Washiashi Masashi Odashima Keiichirou Nemoto
- 出版者
- Japanese Society for Plant Biotechnology
- 雑誌
- Plant Biotechnology (ISSN:13424580)
- 巻号頁・発行日
- vol.40, no.3, pp.229-236, 2023-09-25 (Released:2023-09-25)
- 参考文献数
- 39
- 被引用文献数
- 3
Japanese cultivated gentians are highly valued ornamental flowers in Japan, but the flower shape is mostly limited to the single-flower type, unlike other flowers such as roses and carnations. To overcome this limitation, we used the CRISPR/Cas9 genome editing system to increase double-flowered genetic resources in gentians. Our approach targeted an AGAMOUS (AG) floral homeotic gene (AG1), which is responsible for the natural mutation that causes double flowers in gentians. We designed two targets in exon 1 of AG1 for genome editing and found that 9 of 12 herbicide-resistant shoots had biallelic mutations in the target regions of AG1. These nine lines all produced double flowers, with stamens converted into petaloid organs, similar to the natural mutant. We also analyzed the off-target effects of AG2, which is homologous to AG1, and found that such effects occurred in gentian genome editing but with low frequency. Furthermore, we successfully produced transgene-free genome-edited plants (null segregants) by crossing with wild-type pollen. F1 seedlings were subjected to PCR analysis to determine whether foreign DNA sequences, two partial regions of the CaMV35S promoter and Cas9 gene, were present in the genome. As a result, foreign genes were segregated at a 1 : 1 ratio, indicating successful null segregant production. Using PCR analysis, we confirmed that four representative null segregants did not contain transfer DNA. In summary, our study demonstrates that the CRISPR/Cas9 system can efficiently produce double-flowered gentians, and null segregants can also be obtained. These genome-edited plants are valuable genetic resources for future gentian breeding programs.
- 著者
- Dai ISHIYAMA Tomoko MIZOMOTO Chise UEDA Nobuyuki TAKAGI Noriko SHIMIZU Yu MATSUURA Yuto MAKUUCHI Aiko WATANABE Yasunori SHINOZUKA Kazuhiro KAWAI
- 出版者
- JAPANESE SOCIETY OF VETERINARY SCIENCE
- 雑誌
- Journal of Veterinary Medical Science (ISSN:09167250)
- 巻号頁・発行日
- vol.79, no.3, pp.626-631, 2017 (Released:2017-03-23)
- 参考文献数
- 23
- 被引用文献数
- 10 13
The main factors affecting the outcome of Trueperella pyogenes (T. pyogenes) mastitis were examined through a survey of diagnostic data and interviews relating to the occurrence of T. pyogenes mastitis in 83 quarters from 82 Holstein cows between August 2012 and April 2014. Ultimately, one cow was sold during the examination, and 82 quarters from 81 cows were used for analysis on prognosis. T. pyogenes mastitis occurred year round in both lactating and dry cows. The incidence of T. pyogenes mastitis did not significantly differ by month or show seasonality in either lactating or dry cows. Therefore, the occurrence of T. pyogenes mastitis also differed from that of summer mastitis. The 1-month survival rate of infected cows was 64.6% (53/82), and the recovery rate of quarters with T. pyogenes mastitis was 14.6% (12/82). Bivariate logistic regression analysis was performed with survival and culling of infected cows as objective variables and with recovery and non-recovery of quarters with T. pyogenes mastitis as objective variables. The severe cases were significantly culled (odds ratio, 16.30) compared to mild cases, and the status of quarters didn’t recover (odds ratio, 6.50). The results suggest that mild to moderate symptom severity at the time of onset are the main factors affecting outcomes in cows and recovery of quarters infected with T. pyogenes mastitis. Further, high level of NAGase activity also suggested the potential use as an indicator of culling of cows with T. pyogenes mastitis.