| Aim:Studies on the self-incompatibility(SI) have been mostly focused on the area of pistil S gene and its products. Many progresses on clone, expression, separation, purification and function of 5 gene have been achieved. But there is still a long way for mankind to understand the mechanisms of SI throughout. Although it's believed that the interactivity of pistil and pollen results in the SI reaction, the mechanism of signal transduction during the pistil and pollen recognition is less reported. At the same time, laser confocal scanning microscope (LCSM) technique develops rapidly and is applied in almost all fields of biological research. But it's difficult to process the data of LCSM because, generally, the data can only be processed with LCSM system. This restriction makes researchers limited in data analysis time. So a new convenient way of LCSM data processing is also indispensable.Methods:A most cultivated breed Pyrus serotina Rehd. was selected as experimental samples. Some varieties such as Hohsui(genotype is S3S5), Kikusui (S2S4) , Kohsui (S4S5) and Nijisseiki(S2S4) were used to make various pollination combinations. Pollens were labeled with Fluo-3 AM at low temperature. The labeled pollens were pollinated to the pistil or treated with endogenetic or ectogenetic RNases. The treated pollens and styles were studied with BioRAD?MRC 1024ES LCSM equipped with Zeiss lens. The experiment data were processed with personal computer.Results:The methods of processing LCSM data with personal computer (PC) out of LCSM system were given out. With these methods, it's possible and convenient to read the image stages, add or remove psyudo-color, montage, convert and process the Time Course data and so on. To do so, researchers would have much more time to analysis and investigate the LCSM data.A set of pollen fluorescence labeling methods were established. With the methods, pear pollens can be easily labeled and remain alive. Also, methods of processing LCSM data with PC detached from LCSM system were put forward. With these methods, many daily data processing job can be done in your own PC out of laboratory, for example, reading of image stage, adding/removing pseudo color, montage, processing of TimeCourse data and so on.The dynamic distribution of pollen cytosolic free Ca2+ ([Ca2+]j) after self- and cross-pollination in Pyrus serotina Rehd. has been studied with laser confocal scanning microscopy (LCSM). The compatible and incompatible pollination combinations are shown as follows: Hohsui (#) X Kikusui ( @) and Hohsui (#) X Hohsui (@). Hohsui and Kikusui pollen cells used for observation were stained with Fluo-3 AM at 4$, at which the esterase activity within the cell wall of pollen grain was inhibited to a large extent, and thus a better labeling effect was obtained. Data of pollen germination in vitro show that there is an apparent change in [Ca2+]j. Pollen [Ca2+]i is high before germination and drops down to background after germination. It implicates that Ca2+ affects the pollen germination course of P. serotina and is consistent with many other similar researches. Results of semi- in vivo pollination experiments show that the [Ca2+]j gradient near the germination pore of compatible pollen was lost just because there was a general increase of [Ca2+]j throughout the cell, except for the area near the pollen cell membrane. Following that, the [Ca2+]j dropped asymmetrically and the gradient near the germination pore reconstructed, while the incompatible pollens had no such characteristic [Ca2+]j variation.The effects of endogenetic and ectogenetic RNases on the [Ca2+]j variations of pre-germination pollen were tested. The results showed that the [Ca2+]j variations were complex. In 1 hour, ectogenetic RNases, such as RNase TI and RNase A, and endogenetic incompatibile 'Hohsui' RNase promoted the [Ca2+]j of 'Hohsui' pollen. Acid proteins of 'Hohsui' had no influence on the [Ca2+]j of itself pollen. But endogenetic compatibile 'Kohsui' RNase reduced the [Ca2+]j of 'Hohsui' pollen. According t...
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