烟草幼嫩花粉原生质体分离与早期离体发育

ISOLATION AND EARLY IN VITRO DEVELOPMENT OF YOUNG POLLEN PROTOPLASTS IN NICOTIANA TABACUM

建立了两种分离烟草（Ｎｉｃｏｔｉａｎａ ｔａｂａｃｕｍ ）幼嫩花粉原生质体的方法：一为通过花药漂浮培养释放出外壁裂开的花粉，后者转入酶液彻底脱去外壁，内壁降解而分离出原生质体（简称“花药预培养法”）；二为花粉经饥饿处理后，转入酶液释放原生质体（简称“花粉饥饿预处理法”）。对影响分离效果的主要因素作了研究。用花药预培养法分离的幼嫩花粉原生质体，在Ｋ３ 培养基中可再生细胞壁，启动１ 次细胞分裂，或萌发花粉管。表明具有孢子体发育与配子体发育的潜能。观察到花粉原生质体分裂成二细胞后，其中１ 个子细胞又长出花粉管的稀有现象。

For isolating young pollen protoplasts in Nicotiana tabacum. The authors had established two efficient enzymatic methods via anther preculture or pollen starvation pretreatment. Procedure of the first method included the following steps: 1. Cold pretreatment of flower buds with pollen at late unicellular to early bicellular stage; 2. Anther floating culture for pollen shedding into the culture medium followed by dehiscence of exine; 3. Enzymatic maceration of exine dehisced pollen resulting in degradation of intine and release of pollen protoplasts in large quantity. Procedure of the second method involved the following steps: 1. Culture of pollen at middle bicellular in Kyo and HaradaB medium for starvation; 2. Enzymatic maceration of starvated pollen resulting in release of pollen protoplasts and subprotoplasts. Factors affecting the results of both methods as well as early in vitro developmental events of young pollen protoplasts were studied. The protoplasts could be induced either to trigger the first sporophytic division or to continue the gametophytic pathway leading germinatation of pollen tubes indicating their potentiality of inducing both sporophytic and gametophytic development of pathway. In rare instance a quite interesting phenomenon was observed that a pollen protoplast first divided into two daughter cells and one of which then germinated a pollen tube. It may insinuate that such pollen protoplasts initially induced a sporophytic pathway could reverse induce a gametophytic pathway.