烟草雌性细胞原生质体的融合实验

SINGLE PAIR FUSION OF VARIOUS COMBINATIONS BETWEEN FEMALE GAMETOPLASTS AND OTHER PROTOPLASTS IN NICOTIANA TABACUM

将聚乙二醇诱导的单对原生质体融合技术应用于烟草（Ｎｉｃｏｔｉａｎａ ｔａｂａｃｕｍ ）雌性细胞体外融合，成功地进行了雌性细胞间，雌、雄性细胞间，雌性细胞与体细胞间各种组合的融合实验。此外，应用微滴培养与微室饲养培养两种方法诱导了体细胞原生质体分裂并形成细胞团。

This paper reports an enzymatic maceration osmotic shock method for isolation of tobacco embryo sac and its component cell protoplasts, and also a new method for fusion between single pairs of selected mesophyll protoplasts using polyethylene glycol (PEG) as on inducing agent.An integration of these two methods has lead to the successful fusion of female gametoplasts with other kinds of protoplasts.The female gametoplasts described here, in a broad sense, include the egg cell (E), central cell (C) and synergid (S).One of the female gametoplasts was selected and fused with another female, male (generative cell, G) or somatic (mesophyll, M) protoplast.Various combinations were involved: E+S, E+C, E+G, E+M, C+C, C+S, C+G, C+M, S+S, S+G, S+M, etc..Briefly, the authors were able to choose any desired combination to realize single pair fusion by the new PEG method.For the purpose of culturing such fusion products that were limited in number, the authors had done some preliminary experimets using mesophyll protoplasts as feeder cells.Two methods were adopted: the microdrop culture, and the millicell culture with feeder cells.The mesophyll protoplasts were precultured for 2—3 days in large for population expansion before they were used as feeder cells.One or several protoplasts were cultured in a microdrop or a millicell and were induced formation of small cell clusters.This result indi cated that the culture methods might also be suitable for culturing the products from fusion of female gametoplasts and other protoplasts in this plant species.