利用辣椒疫霉培养滤液体外筛选胡椒抗瘟病无性系研究

In vitro Selection of Black Pepper (Piper nigrum Linn.) Somaclones Resistant to Foot Rot Using Culture Filtrate of Phytophthora capsici

摘要:

在胡椒(PipernigrumLinn.)茎尖丛生增殖技术的基础上,以印尼大叶种“LampongType”无菌实生苗作外植体源,利用辣椒疫霉(Phytophthoracapsici)培养滤液对胡椒茎尖及其增殖形成的丛生芽进行体外选择。辣椒疫霉培养滤液的不同灭菌方法对辣椒疫霉培养滤液的毒性影响显著,过滤灭菌方式可以保持辣椒疫霉培养滤液的毒性,而高温高压灭菌方式则不能。随着辣椒疫霉培养滤液浓度的增加,茎尖和丛生芽的存活率和增殖率都在下降。在存活的茎尖或丛生芽培养中,一部分可正常增殖,其余的形成愈伤组织,或者保持生长停滞的休眠状态。在选择性培养基上继代培养2次后进行生根和移栽,利用离体叶片针刺接种法对温室条件下生长的移栽植株进行抗瘟病测定。以3次抗病检测均无明显症状的植株作为抗病株。随着辣椒疫霉培养滤液浓度的增加,得到的再生植株数量降低,但其中抗病株的比例提高。利用过滤灭菌方式加入选择性培养基的处理中,25%、50%和75%的辣椒疫霉培养滤液分别获得1株、4株和3株抗病株,分别占各处理再生植株总数的1.54%、20.00%和42.86%,共获得8株,占该组处理再生植株总数的8.70%。

Abstract:

Based on shoot-tip multiplication technique, the application of in vitro selection of black peppersomaclones resistant to foot rot disease caused by fungus Phytophthora capsici was carried out using a large-leaf variety Daye (Lampong Type) which is widely cultivated in Hainan but highly susceptible to P. capsici as explant resources. The results demonstrated that sterilization methods significantly influenced on the toxicity of the culture filtrate of P. capsici. The fungal filtrate toxicity could be maintained using filtrate sterilization by addition of the fungal filtrate into selective medium instead of autoclave sterilization． The survival rate of shoot tips and multiple shoots decreased with the increasing concentrations of the fungal culture filtrate． Some cultures blackened and died finally, and some formed calli or remained dormant．After 2 subcultures onto the same selective medium， microshoots were rooted in vitro and tran sferred to the greenhouse conditions for screening the resistance to fungus P. capsici．The plants exhibited no external symptoms of the disease during three successive times of resistance assay once two weeks were considered to be resistant． As the concentrations of fungal culture filtrate increased，the total number of regenerated plants obtained declined but the frequencies of plants resistant to capsici increased．1(1.54％)，4(20.OO％) and 3(42.86％) disease resistant somaclonal variants were obtained at concentration of 25％ ，50％ and 75％ fungal culture filtrate as selective agents， respectively，with the total number of 8 plants resistant to P. capsici.