杨树2n花粉花粉管生长缓慢的细胞学机理

Triploid poplars play important roles in landscape cultivation, ecological protection and the production of lumber and pulp in China. It is one of vital methods to obtain triploids using 2n pollen crossing normal female gamete, which combined the ploidy effect with heterozygosity. There existes competition between 2n pollen grains and normal pollen grains as 2n pollen can not be screened from mixed pollen. However, pollen tubes slowly grow on stigma when pollinated with mixed pollen grains resulting in a lower frequency of triploid due to their poor competition. However, little is known about the cytological mechanism of slow polar growth of pollen tube in 2n pollen. In this study, we focused on the polar growth of 2n pollen during fertilization. Based on the investigation of germination rates of normal pollen, natural 2n pollen and induced 2n pollen and calculation of growth rates of their pollen tubes in vitro, the effects of ultrastructural observation, actin microfilament, Calcium signals and its distribution, the Endo/ Exocytosis to polar growth of pollen tube will be conducted to reveal the cytological mechanism of poor competition of 2n pollen during fertilization using natural 2n pollen producers of Populus tomentosa as materials. All results will provide a theoretical platform for triploid breeding of poplar.

利用2n 花粉与单倍性雌配子杂交是杨树三倍体选育的一条重要途径，实现了花粉染色体加倍带来的倍性效应与杂交产生的杂种优势的结合。在2n花粉利用时，由于2n花粉与单倍性花粉难以分离纯化，授粉时存在单倍性花粉竞争，2n花粉的花粉管生长缓慢而致使三倍体得率低。但2n花粉的花粉管生长缓慢的细胞学机理至今仍不清楚。本研究围绕杨树2n花粉花粉管生长缓慢的科学问题，以天然2n花粉高产的毛白杨为研究材料，在研究单倍性花粉、天然2n花粉以及人工诱导2n花粉三种类型花粉在离体萌发条件下萌发率和花粉管生长速率差异的基础上，研究其花粉管超微结构、微丝骨架的排列方式与动态组装调控、Ca2+浓度剃度及分布、胞吞胞吐作用等对花粉管极性生长的影响，进而揭示2n花粉花粉管生长缓慢的细胞学机制，并采取适宜的技术措施，提高杨树三倍体得率。

针对杨树2n花粉花粉管生长缓慢导致三倍体得率的科学问题，以毛白杨无性系5088和胡杨雄花枝为材料，在研究毛白杨以及胡杨小孢子发生发育、天然2n花粉发生机理、2n花粉的形态学识别标准、2n花粉的人工诱导以及适宜离体萌发培养基筛选的基础上，比较分析了LATB处理单倍性花粉以及2n花粉萌发率、花粉管生长速率、花粉管壁物质积累的差异，并就其花粉管胞吞胞吐、超微结构、微丝骨架的排列、Ca2+浓度梯度与花粉管极性生长关系进行了研究。发现减数分裂后期Ⅱ，纺锤体微管缺失致使姊妹染色体单体分离后不移向两极是SDR型2n花粉形成的一种重要机制；提出了以无性系花粉粒平均直径的1.28倍作为2n花粉形态判别的动态标准；筛选出12％蔗糖+50mg/L氯化钙+120mg/L硼酸+0.7％琼脂为毛白杨花粉离体萌发的适宜培养基。明确了LATB处理对单倍性花粉及2n花粉萌发率、花粉管生长速率、花粉管壁物质积累的影响规律，单倍性花粉管生长速率、花粉管壁体积均远远大于2n花粉管，但2n花粉管壁厚度较单倍性花粉管大。揭示了2n花粉管内细胞器结构异常且数量偏少，高尔基体片层及线粒体膜结构遭到一定程度破坏，造成合成细胞壁前体物质的速率较单倍性花粉管低；同时，其钙离子浓度梯度不明显，微丝骨架异常，呈现垂直于生长轴的方向，使高尔基体分泌囊泡定向运输与融合不能够正常进行或速率降低是2n花粉花粉管生长缓慢的细胞学机制。