摘要
以木本植物杨树(Populus sp.)和核桃(Juglans regia L.)为材料,对内源生长素免疫胶体金定位技术在固定、烤片、免疫染色、显色等关键环节进行了改进优化与验证.结果显示,优化后适合木本植物定位方法的主要技术要点是:在染色中,通过采用尿素-胰蛋白酶联合消化技术和增加牛血清白蛋白处理大大地改善了抗原修复结果,提高了染色的敏感性和特异性.利用优化后的方法对核桃幼胚和杨树嫩茎诱导生根过程中的吲哚乙酸(IAA)进行定位研究,发现杨树试管嫩茎生根过程中,形成层及周缘维管束有很强的IAA信号,核桃子叶生根中,胚中有很强的IAA信号,胚根中有半圆形、胚芽中有>"形强信号区,胚轴信号较弱,胚根信号最强.研究表明,与传统免疫染色方法相比,优化后的方法对木本植物生长素定位具有敏感性高,特异性强,银颗粒明显,背景清晰,耗时少等特点.
In this research,the technique of immuno-gold localization of endogenous auxin was investigated with walnut and poplar. The main and key steps such as fixation,dying slides,immunostaining were improved. The results show that the main points of the improved methods which is suitable for woody plants were urea together with trypsin treatment and BSA （bovine serum albumin） added in digestion in order to unmask the antigens of tissue sections and increase the sensitivity. This procedure was successfully applied to localize the distribution of IAA （indole-3-acetic acid） in walnut embryo and during rhizogenesis of poplar shoots. The results showed that during the rhizogenesis of poplar tender shoots,IAA was mainly distributed in cambium and the vascular bundle and in the embryo of walnut, the high density of silver particles was found in radicle and plumule,the signal in radicle was semi-circularity shape and in plumule was ＂〉＂ shape, The research reveals that compared with traditional procedure of immuno-staining, the improved methods have characters of high sensitivity,strong speciality,obvious silver paricles and clear backgroud.
出处
《西北植物学报》
CAS
CSCD
北大核心
2008年第4期819-825,共7页
Acta Botanica Boreali-Occidentalia Sinica
基金
国家自然科学基金项目(30671436)资助
