Improvement of anther cultures conditions using the Taguchi method in three cereal crops

Background: Plant tissue cultures have the potential to reprogram the development of microspores from normal gametophytic to sporophytic pathway resulting in the formation of androgenic embryos. The efficiency of this process depends on the genotype, media composition and external conditions. However, this process frequently results in the regeneration of albino instead of green plants. Successful regeneration of green plants is affected by the concentration of copper sulfate (CuSO4) and silver nitrate (AgNO3) and the length of induction step. In this study, we aimed at concurrent optimization of these three factors in barley (Hordeum vulgare L.), wheat (Triticum aestivum L.), and triticale (x Triticosecale spp. Wittmack ex A. Camus 1927) using the Taguchi method. We evaluated uniform donor plants under varying experimental conditions of in vitro anther culture using the Taguchi approach, and verified the optimized conditions. Results: Optimization of the regeneration conditions resulted in an increase in the number of green regenerants compared with the control. Statistic Taguchi method for optimization of the in vitro tissue culture plant regeneration via anther cultures allowed reduction of the number of experimental designs from 27 needed if full factorial analysis is used to 9. With the increase in the number of green regenerants, the number of spontaneous doubled haploids decreased. Moreover, in barley and triticale, the number of albino regenerants was reduced. Conclusion: The statistic Taguchi approach could be successfully used for various factors (here components of induction media, time of incubation on induction media) at a one time, that may impact on cereals anther cultures to improve the regeneration efficiency

Downregulation of OsAGO17 by artificial microRNA causes pollen abortion resulting in the reduction of grain yield in rice

Background: Pollen development is an important reproductive process that directly affects pollen fertility and grain yield in rice. Argonaute (AGO) proteins, the core effectors of RNA-mediated silencing, play important roles in regulating plant growth and development. However, few AGO proteins in rice were reported to be involved in pollen development. In this study, artificial microRNA technology was used to assess the function of OsAGO17 in pollen development. Results: In this study, OsAGO17, a rice-specific gene, was specifically expressed in rice pollen grains, with the highest expression in uninucleate microspores. Downregulation of OsAGO17 by artificial microRNA technology based on the endogenous osa-miRNA319a precursor was successfully achieved. It is found that downregulation of OsAGO17 could significantly affect pollen fertility and cause pollen abortion, thus suggesting that OsAGO17 functions in rice pollen development. In addition, the downregulation of OsAGO17 mainly caused a low seed-setting rate, thereby resulting in the reduction of grain yield, whereas the downregulation of OsAGO17 did not significantly affect rice vegetative growth and other agricultural traits including number of florets per panicle, number of primary branch per panicle, and 100-grain weight. Furthermore, the result of subcellular localization analysis indicated that the OsAGO17 protein was localized to both the nucleus and the cytoplasm. Conclusion: These results represent the first report of the biological function for OsAGO17 in rice and indicate that OsAGO17 may possibly play crucial regulatory roles in rice pollen development. It helps us to better understand the mechanism of pollen development in rice.