激光、高压静电场克服羽衣甘蓝自交不亲和及其分子机制的研究

Overcoming Self-incompatibility of Brassica oleracea var.acephala by Laser and High-voltage Electrostatic Field and Molecular Mechanism

利用激光、高压静电场对自交不亲和的羽衣甘蓝的花粉进行处理,以期克服其自交不亲和。通过对处理后的花粉的表面结构和蛋白酶活性及其萌发情况的研究,进一步分析激光、高压静电场的生物效应。同时利用分子生物学手段Northern杂交技术,检测了经激光、高压静电场处理的花粉,在授粉后柱头上SLG和SRK基因表达的差异,以探讨激光、高压静电场对生殖反应作用的分子机制。结果表明:通过适宜剂量的激光、高压静电场处理,羽衣甘蓝的花粉萌发率提高,成熟花粉水溶性总蛋白,α-淀粉酶活力,总淀粉酶活力均不同程度的提高,同时自交亲和指数和结籽率提高。从花粉的表面结构的电镜显微观察分析,花粉壁的雕纹受到了伤害,网脊线有轻微断裂,网眼变大。授粉后,取不同时间间隔的柱头样品提取总RNA,利用Northern杂交技术检测SLG和SRK基因的表达量,授粉后24 h激光处理和高压静电场处理与对照比较,SLG和SRK基因的表达量均降低。

Incompatible cultivars of Brasstca oleracea var. acephala were used as test materials in this experiment. The test materials were treated by laser and high-voltage electrostatic field for overcoming their self-incompatibility. The biological effect of laser and high-voltage electrostatic field was studied further by analyzing the surface structure, proteinase activity and germination for the treated pollen. The action mechanism of laser and high-voltage electrostatic field to generative reaction was discussed by analyzing expression difference of SLG and SRK after pollination of the treated pollen by Northern blotting. The results are as follows： （ 1 ） Through treatment in appropriate dosage of laser and high-voltage electrostatic field, the budding quantity of pollen of Brassica oleraeea var. acephala was increased, the self-compatibility index and the quantity of seed were increased. （2） The surface structure of the pollen was observed by electron microscope. It was found that the sculpture of the pollen treated by laser and high-voltage electrostatic field was hurt, some reticulate was broken and the net hole enlarged. The concentration of protein, activity of α-amylase and amylase in the pollen was improved. （3） After self pollination by the pollen treated by laser and high-voltage electrostatic field, total RNA was extracted from stigma tissue picked in different time interval pistils, and the expression of ＇SLG SRK＇ was tested by Northern blotting. The results showed that the expression of ＇ SLG SRK＇ of the laser and high-vohage electro- static field treated materials was lower than that of control after 24 hours of pollination.