Studies On The Programmed Cell Death During Mutant Short Catkin Development In Chestnut

One important aim in chestnut breeding is to select the high yield and the large ratio of female flowers to male inflorescence. The mutant short catkin chestnut is a rare germplasm resource, which was discovered in Miyun district of Beijing. The mutant chestnut has the remarkable small short catkin, and during the catkins development, the upper parts of the catkins appear natural death gradually, while the remainder catkins still developed normally. The physiological mechanism of naturally shorten on the mutant catkins was studied by biological observation, paraffin slices, electron-transmission microscope, DNA Ladder analysis and TUNEL assay, the results are as follows:1. The differentiation pattern of mutant chestnut male flowerThe male flower differentiation lasted for about 10 months, beginning May-June and ending April-May of the following year, The differentiation process could be divided into 6 stages, from formation of the primordium of the male inflorescence, to the formation of anther, that was consistent to the contrast; There was a special phenomenon, the male flower developed only to the floral envelop differentiation in the upper catkin, and the cells happened abnormal death ,while the remainder catkin still developed normally; it showed that, the abnormal cell death induced the degradation of upper catkins, and shortened the catkin.2. The ultrastructure changes of cells during the upper catkins naturally degradationThe ultrastructure changes of abnormal cell death in upper catkins were observed by using electron-transmission microscope, the nucleus showed chromatin condensation, the nucleolus dissolved, karyoplasm degraded; the karyotheca burst and the nucleus disintegrated; the vacuole happened distinct swallow, and some formed into many small vacuoles; the chloroplast and mitochondrion disintegrated gradually; The morphological changes during the death of all the organelles within the cell showed the typical features of programmed cell death, so it indicated that the abnormal cell death was induced by programmed cell death.3. Detection of fragmented nuclear DNA in the short catkinIn this work, the fragmented nuclear DNA in the short catkin was investigated by using microscopy, total DNA agarose gel electrophoresis and TUNEL assay. Agarose gel analysis of total DNA showed the DNA Ladder, and the TUNEL-positive nuclei were detected in the cells of upper short catkin, indicating the total DNA was ruptured into fragmentations. Therefore, it was proved that the programmed cell death had happened in the upper catkin, while it development.