Development Of Techniques Of Assessment On Biosafety Of Gene Flow Of Transgenic Herbicde-Resistant Rice Based On Sexual Compatibility

The main problem to release transgenic plant in the field is the target-gene flow to wild plant and causes potential danger to farmland ecological environment. So it is essential to make a precise assessment about gene flow before transgenic plant field release.Sexual compatibility between species in natural conditions plays a key role in occurrence of gene flow. The methods of assessment of occurrence stage of sexual incompatibility between species were developed on the basis of sexual compatibility between species by means of reproductive biology. The objectives of the studies are to develop assessment techniques of possibility of gene flow through observing sexual compatibility at the early stage using trans-genic rice and weedy rice, medical wild rice (Oryza officinalis) and Echinochloa spp. as experimental materials, and ultimately to integrate those techniques into safety assessment system of transgenic crops at individual and population level. The main content is given as follows:Hybridization incompatibility may occur at many stages in reproduction. It includes pollens can't germinated and pollen tubes can't enter the stigma or reach the ovary, the fruit or the seed can't develop, hybrid seeds die, sterile or damage. Looking for the simple, quick methods to observe incompatibility stage, especially at very early stage, can offer strong support for the assessment. This is also the weak link on environmental safety assessment research and technique at present comparatively.The first stage, whether pollen can germinate and pollen tube enter the stigma, could be clearly observed under ordinary microscope. Pollen tubes of the transgenic rice pollinated on the stigma of barnyard grass (Echinochloa spp.) were so twisted and out of shape that can't enter the stigma. Therefore they couldn't hybrid and the possibility of gene flow should be extremely low.It is difficult to observe the second stage of incompatibility, namely pollen tube can't enter the ovary with the above ordinary microscope, because the pollen tube is growing inside the style tissue. Pollen tube stained with the peculiar fluorescence dyestuffs could present bright yellowish green in style, and ovary be observed under the fluorescence microscope, while the style and ovary of spontaneous weak fluorescence are distinguished with very dark color comparing the stained pollen tubes. The above method for judge thesecond stage was convenient. When the transgenic rice pollen pollinated on the medical wild rice stigma was observed with this method, it was found that the pollen could germinated and entered the stigma normally, but the tube expanded and stopped growth when reaching the style base. This proved that hereditary materials of the transgenic rice could reach the medical wild rice style, so the possibility of gene exchange might increase. From above, it was concluded that possibility of gene flow from transgenic rice to the medical wild rice was relatively low but higher than to barnyard grass.Third stage, namely fruit or seed can't develop, hybrid seed die, could be judged by observing the embryo sac development mainly. After cutting ovary buried with paraffin into 8 microns slices and dyed, the structure of the embryo sac including in the slices was observed under ordinary microscope accurately, clearly, and meticulously. Transgenic rice pollen on weedy rice's stigma was observed with the above technology. The results showed that transgenic rice pollen could enter the embryo sac of weedy rice normally, but seed rate were low. It was found that embryo and endosperm development was incongruity although fertilization could occur. From above it was concluded that the possibility of gene flow was relatively high.But operation course of paraffin quite was great superfluous, and setting up three-dimensional concept according to plan slices observation needed patience and experience.The whole clearing method was to observe embryo directly after dealing with wholly ovary by transparent, which had advantages of easy and simple handling and three-dimensional eff...