三种贝母（Fritillaria spp.）的快繁及离体保存技术研究
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摘要
本文以三种观赏贝母—皇冠贝母( Fritillaria imperialis )、弯尖贝母(F.acmopetala)、波斯贝母(F.persica)为材料,进行了组培快繁,组培苗生根结鳞茎培养及常温抑制保存技术研究。为建立观赏贝母的快繁技术体系、种质资源的安全和有效保存及观赏贝母的推广应用提供技术依据。主要结果如下:
1.观赏贝母无菌无性系的建立
观赏贝母的鳞茎先以10%NaClO消毒20min,然后再用0.1%HgCl2消毒10min效果最好,而叶片、花丝、鳞芽等较干净部位选用10%NaClO消毒18～20min;用三种贝母的鳞茎、带基盘鳞片、花丝、鳞芽及叶片进行培养,以带基盘鳞片和花丝诱导分化效果最理想;用皇冠贝母的无菌苗叶片作为外植体,取得了良好的分化效果。
皇冠贝母鳞茎初代最适合诱导培养基为MS+BA1.0mg/L+NAAO.2mg/L,弯尖贝母为MS+BA1.0mg/L+NAA0.1mg/L,皇冠贝母无菌苗叶片为MS+BA 2.0mg/L+NAA0.5mg/L。而波斯贝母在各培养基组合中培养效果均不理想,其适合的培养条件有待于进一步研究。
2.继代增殖—大量繁殖
皇冠贝母和弯尖贝母均以培养基MS+BA0.5mg/L+NAA0.1mg/L的增殖率最高,增加苗数最多。
3.生根结鳞茎
降低大量元素的浓度有利于生根结鳞茎,较适的浓度为1/4MS;NAA有利于根的生长,不利于鳞茎的形成,糖也有助于鳞茎的形成,较适浓度为60g/L;培养基中添加活性炭以及一定时间的黑暗培养也有利于鳞茎生长。
4.炼苗移栽
弯尖贝母组培苗最佳的移栽介质为泥炭土∶砻糠灰∶珍珠岩=7:2:1,成活率能达到100%。
5.种质离体保存
在MS或者大量微量元素均减半的1/2MS培养基添加1.0～3.0mg/LABA;70～90g/L高浓度蔗糖能够保存弯尖贝母试管苗9个月以上。而10～5Og/L甘露醇的抑制效果不是太明显。
In this experiment,effective multiplication method,rooting and bulblet formation and germplasm conservation in vitro were studied based on onamental Fritillaria of three varieties-F.imperialis,F.acmopetala,F.persica.In order to establish the propagation system for appliance and extend of ornamental fritillarias.The main results are as followings:
1.Effective multiplication method was set up.
In the period found of axenic clone 10%NaClO and 0.1% HgCl2 is the best disinfector to deal with the bulb,as to neat part, such as leave, filament, and bulbil is fit to use 10%Naclo 18～20mins.And found the best effect is the bulb scale with base and filament,leaf was not inducement adventitious buds.Free leaf of F. imperialis is induced better.
F.imperialis is fit MS+BA1.0mg/L+NAA0.2mg/L,F.acmopetala is fit MS+ BA1.0mg/L+NAA0.1mg/L,free leaf of F.imperialis is fit MS+BA2.0mg/L+NAA0.5mg/L,but the medi-um fit to culture F.persica haven't been found.
2. In the period of Subculture-Found of mass production,the most multiplication of adventitious buds in MS+BA0.5mg/L+NAA 0.1mg/L during subculture of F.imperialis and F.acmopetala.
3. Stage of rooting and bulblet formation.
It was favorable for rooting and bulblet formation of F.acmopetala in reducing the concentration of macroelement.1/4MS proved the best concentration.NAA promoted rooting but made against forming bulblet.Sucrose can help to form bulblet, the best concentration was 60g/L.
4. Transplanting.
Transplanting substrate with Peat:Longkang Gray:perlite=7:2:1 was the best,where the survival rate could reach 100%.
5.The conservation of Fritillaria plantlets.
The results showed that,MS or half strength MS medium with 1.0～3.0mg/L ABA;the MS medium with 70～90g/L sucrose restricted in vitro growth of the shoots efficiently and prolonged the interval between subcultures over 9 months.Otherwise,10～50g/L mannitol could not restrict the growth of in vitro shoots effectively.
1.观赏贝母无菌无性系的建立
观赏贝母的鳞茎先以10%NaClO消毒20min,然后再用0.1%HgCl2消毒10min效果最好,而叶片、花丝、鳞芽等较干净部位选用10%NaClO消毒18～20min;用三种贝母的鳞茎、带基盘鳞片、花丝、鳞芽及叶片进行培养,以带基盘鳞片和花丝诱导分化效果最理想;用皇冠贝母的无菌苗叶片作为外植体,取得了良好的分化效果。
皇冠贝母鳞茎初代最适合诱导培养基为MS+BA1.0mg/L+NAAO.2mg/L,弯尖贝母为MS+BA1.0mg/L+NAA0.1mg/L,皇冠贝母无菌苗叶片为MS+BA 2.0mg/L+NAA0.5mg/L。而波斯贝母在各培养基组合中培养效果均不理想,其适合的培养条件有待于进一步研究。
2.继代增殖—大量繁殖
皇冠贝母和弯尖贝母均以培养基MS+BA0.5mg/L+NAA0.1mg/L的增殖率最高,增加苗数最多。
3.生根结鳞茎
降低大量元素的浓度有利于生根结鳞茎,较适的浓度为1/4MS;NAA有利于根的生长,不利于鳞茎的形成,糖也有助于鳞茎的形成,较适浓度为60g/L;培养基中添加活性炭以及一定时间的黑暗培养也有利于鳞茎生长。
4.炼苗移栽
弯尖贝母组培苗最佳的移栽介质为泥炭土∶砻糠灰∶珍珠岩=7:2:1,成活率能达到100%。
5.种质离体保存
在MS或者大量微量元素均减半的1/2MS培养基添加1.0～3.0mg/LABA;70～90g/L高浓度蔗糖能够保存弯尖贝母试管苗9个月以上。而10～5Og/L甘露醇的抑制效果不是太明显。
In this experiment,effective multiplication method,rooting and bulblet formation and germplasm conservation in vitro were studied based on onamental Fritillaria of three varieties-F.imperialis,F.acmopetala,F.persica.In order to establish the propagation system for appliance and extend of ornamental fritillarias.The main results are as followings:
1.Effective multiplication method was set up.
In the period found of axenic clone 10%NaClO and 0.1% HgCl2 is the best disinfector to deal with the bulb,as to neat part, such as leave, filament, and bulbil is fit to use 10%Naclo 18～20mins.And found the best effect is the bulb scale with base and filament,leaf was not inducement adventitious buds.Free leaf of F. imperialis is induced better.
F.imperialis is fit MS+BA1.0mg/L+NAA0.2mg/L,F.acmopetala is fit MS+ BA1.0mg/L+NAA0.1mg/L,free leaf of F.imperialis is fit MS+BA2.0mg/L+NAA0.5mg/L,but the medi-um fit to culture F.persica haven't been found.
2. In the period of Subculture-Found of mass production,the most multiplication of adventitious buds in MS+BA0.5mg/L+NAA 0.1mg/L during subculture of F.imperialis and F.acmopetala.
3. Stage of rooting and bulblet formation.
It was favorable for rooting and bulblet formation of F.acmopetala in reducing the concentration of macroelement.1/4MS proved the best concentration.NAA promoted rooting but made against forming bulblet.Sucrose can help to form bulblet, the best concentration was 60g/L.
4. Transplanting.
Transplanting substrate with Peat:Longkang Gray:perlite=7:2:1 was the best,where the survival rate could reach 100%.
5.The conservation of Fritillaria plantlets.
The results showed that,MS or half strength MS medium with 1.0～3.0mg/L ABA;the MS medium with 70～90g/L sucrose restricted in vitro growth of the shoots efficiently and prolonged the interval between subcultures over 9 months.Otherwise,10～50g/L mannitol could not restrict the growth of in vitro shoots effectively.
引文
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