高温诱导杨树产生2n花粉的遗传学机制

Triploid breeding which combined the ploidy effect with heterozygosity is one of the most powerful approaches for improvement of the genus Populus L. Growth, wood property traits and adaptability will be improved by only a breeding cycle. In previous studies, high temperature exposure was reported as an effective physical mutagenic agent for inducing of polyploidy plants. It was widely used to obtain triploids and tetraploids via inducing chromosome doubling of megaspore, embryo sac, zygote and pollen. A number of triploids and tetraploids were achieved in Populus. Cytological mechanisms of spontaneous 2n gamete formation were documented in numerous plant species. These mechanisms are currently subdivided as first division restitution (FDR), second division restitution (SDR), post-meiotic restitution (PMR) according to the genetic institution of 2n gametes. To gain Populus triploids, a technology system of inducing 2n gametes by high temperature exposure is built, and it can meet the requirement of triploid production in breeding practice. However, little information on why 2n gametes are produced by High temperature exposure has been known. In the present study, we focused on how pollen mother cells (PMCs) produce 2n gametes after male branches were shocked by high temperature using Populus tomentosa or Populus canescens as materials. The relationships between 2n pollen and chromosome beheaviors, and the microtubule cytoskeleton, and abnormal cytokinesis, and cell plate formation, and genes expression related to meiosis and cytokinesis were investigated. Then genetic mechanism of 2n formation by high temperature exposure would be revealed in Populus. All results will provide a theoretical platform for triploid breeding of Populus.

三倍体育种综合了倍性效应与杂种优势，通过一次育种过程可实现生长、材性以及适应性等多目标性状遗传改良，是杨树遗传改良的重要途径与方法。高温作为一种物理诱变剂，已成功应用于诱导杨树大孢子、胚囊、合子以及花粉染色体加倍选育多倍体的育种实践，获得了大量的杨树三倍体和四倍。有关植物自发产生2n配子机制研究的报导很多，根据配子遗传组成，可分为FDR、SDR以及PMR三种机制。尽管高温诱导杨树2n配子选育三倍体的技术体系业已成熟，完全可以满足三倍体育种的需要，但高温诱导杨树产生2n配子的机制至今仍不清楚。本研究以毛白杨或银灰杨雄花枝为试验材料，以未处理花芽为对照，研究高温处理对杨树花粉母细胞减数分裂染色体行为、微管骨架动态变化，胞质分裂和辐射状微管定位异常，纺锤体微管蛋白以及胞质分裂等相关基因差异表达对2n花粉形成的影响，进而揭示高温诱导杨树2n花粉的遗传学机理，为杨树三倍体育种奠定理论基础。

针对高温处理诱导杨树产生2n花粉的遗传学机制问题，以育性良好的银灰杨为材料，在开展银灰杨雄花芽外部形态与花粉母细胞减数分裂进程对应关系、高温处理诱导银灰杨产生2n花粉技术条件优化的基础上，综合细胞遗传学、分子遗传学以及多倍体诱导技术，系统地开展了高温处理对银灰杨花粉母细胞减数分裂染色体行为、微管骨架动态变化、减数分裂以及胞质分裂相关基因差异表达的影响研究。明确了高温诱导银灰杨花粉染色体加倍的适宜温度、有效处理时期以及持续处理时间，筛选出利用38℃或41℃的高温对发育至双线期至终变期的花粉母细胞进行6h持续处理是高温诱导银灰杨花粉染色体加倍的适宜处理组合。发现了高温处理不但可以诱导银灰杨产生高频率的2n花粉，导致银灰杨花粉产量显著降低，败育花粉比例的提高，还会致使高温诱导型2n花粉表面产生明显的裂沟，但不会影响2n花粉的萌发率的现象。揭示了高温处理诱导银灰杨花粉母细胞周期蛋白A基因CCNA显著下调表达，致使花粉母细胞第二次减数分裂缺失，导致大量二分体的形成，进而发育形成2n花粉可能是高温处理诱导银灰杨产生2n花粉的细胞及分子机制。研究结果对于林木遗传学和三倍体育种基础理论的发展具有重要意义。