In vitro culture and identification of polyembryos from polyembryonic seeds in Manju x Yanxiwanlu Ponkan by leaf morphology and SSR analysis

Abstract：Following the first report of high ratio polyembryonic seeds in monoembryonic Kiyomi x polyembryonic Ponkan, polyembryonic seeds were found in the cross monoembryonic Manju （Citrus tardiferax Hort. ex Tan） x polyembryonie Yanxiwanlu Ponkan （C. reticulata Blanco） also. The ratio of polyembryonic seeds was about 30% generally. To investigate the origin of polyembryos, the embryos from polyembryonie seeds were cultured in vitro. Polyembryonic seedlings from one seed were selected and analysed by leaf morphology and SSR markers. The result of leaf morphology analysis showed that most of the polyembryonic seedlings were significantly different to both parents and indicated zygotic origins, but three seedlings in the 5 seedlings group were similar to the seed parent and it is difficult to determine their origins. Nine specific SSR markers from different citrus linkage groups were screened and used in SSR analysis. Four or more markers were found to show different band patterns among all polyembronic seedlings and seed parent. And it indicated that all seedlings were most likely to be sexual origins. Some of the polyembronic seedlings from one seed were identical to all tested markers and others were different in more than one marker. And this might indicate their different zygotic embryos origins, splitting of zygotic embryos or fertilization of more than one egg.