葡萄柚种子无菌苗组培快繁体系的建立

Establishment of rapid propagation for sterile seedlings of grapefruit by tissue culture

关键词:  葡萄柚  种子  植物生长调节剂  初代培养  继代培养

Key words:  grapefruit  seed  plant growth regulator  primary culture  subculture

摘要:

以葡萄柚种子在无菌条件下萌发的幼苗作为外植体供体,子叶切断作为丛生芽初代诱导培养的接种材料,以MT作为基本培养基,通过调节不同植物生长调节剂及其浓度组合、不同质量浓度的蔗糖,选择最佳初代培养基、继代培养基和生根培养基,对葡萄柚组培快繁技术体系进行研究。结果表明:(1)葡萄柚种子经预处理后,先用75%酒精表面灭菌15 s,再用0.1% HgCl₂浸泡20 min的消毒效果最好,污染率为18.33%,萌发率为89.91%;(2)初代培养的最适培养基为MT+1.0 mg·L^-16-BA(6-苄氨基腺嘌呤)+0.2 mg·L^-1IBA(吲哚丁酸)+0.2 mg·L^-1GA(赤霉素)+蔗糖40 g·L^-1,腋芽诱导率较高,为93.33%;(3)浓度为0.2 mg·L^-1的GA能提高初代培养的腋芽诱导率,但还达不到显著水平;(4)继代培养的最适培养基为MT+0.5 mg·L^-16-BA+0.2 mg·L^-1IBA+0.2 mg·L^-1GA+蔗糖40 g·L^-1,丛生芽增殖系数达到4.25,芽深绿色,茎粗壮,节间较长,生长旺盛;(5)浓度为0.2 mg·L^-1的GA能显著提高继代培养的增殖系数;(6)最适合继代培养的蔗糖浓度为40 g·L^-1,丛生芽长势极好;(7)最佳生根培养基为1/2 MT+NAA 0.2 mg/L+蔗糖40 g·L^-1+AC 0.1%,生根率达68.89%,根粗壮;(8)生根苗移栽30 d后成活率在75%以上。该研究建立了葡萄柚的组织培养快繁技术体系,为葡萄柚的规模化生产提供了可行的技术依据。

Abstract:

The most appropriate sterilization method for tissue culture of grapefruit's seeds was selected, the seedlings sprouted from grapefruit's seeds under sterile conditions acted as explant donors, the cotyledon cuts were used as the inoculation material of the clustered shoots' primary induction culture, the best primary culture medium, subculture culture medium and rooting medium were selected by adjusting the concentrations of sucrose, combinations of different plant growth regulators and their concentrations in the MT medium, and the tissue culture and rapid propagation technique system of grapefruit were studied. The results were as follows:(1)Rapidly immersed with 75% alcohol for 15 s plus 0.1% HgCl₂ for 20 min after the pretreatment was the best disinfection method for grapefruit seeds with the contamination rate of 18.33%, and the germination rate of 89.91%;(2)The best medium for the primary culture was MT+1.0 mg·L^-16-BA(6-Benzylaminopurine)+0.2 mg·L^-1IBA(Indole-3-Butytric acid)+0.2 mg·L^-1GA(Gibberellic acid)+sucrose 40 g·L^-1, the axillary shoot induction rate was up to 93.33%;(3)GA the concentration of which was 0.2 mg·L^-1 could improve the axillary shoot induction rate of primary culture, but couldn't reach significant level;(4)MT+0.5 mg·L^-16-BA+0.2 mg·L^-1IBA+0.2 mg·L^-1GA+sucrose 40 g·L^-1 was the most suitable for the subculture with the multiplication coefficient of 4.25, the shoots were dark green with vigorous growth, the stems were thick and internodes were relatively long;(5)GA the concentration of which was 0.2 mg·L^-1 could significantly improve the multiplication coefficient of subculture;(6)The most suitable concentration of sucrose for the subculture was 40 g·L^-1, the clustered shoots' growth was excellent;(7)The best rooting medium was 1/2MT+NAA 0.2 mg·L^-1+sucrose 40 g·L^-1+AC 0.1%+sucrose 40 g·L^-1 with the rooting rate of 68.89%, the roots were very sturdy;(8)The transplant survival rate of rooting plantlets was up to 75% after being transplanted for 30 d. Through the experiment the tissue culture and rapid propagation technique system of grapefruit was established, which would provided a feasible technical basis for the mass production of grapefruit.