摘要
安祖花是我国近些年引进的一种名贵花卉植物,其花型奇特、花期持久、色彩丰富,深受人们青睐。但以传统的分株方法繁殖安祖花不仅繁殖速度慢、繁殖率极低,而且还受到季节的限制,这些因素极大地限制了安祖花的发展。为了解决这些问题,前人尝试了采用植物离体培养技术来快速繁殖安祖花,遗憾的是目前国内已报道的安祖花组培技术还未能达到工厂化培养的要求,特别是炼苗和移栽技术未能过关,另外,组培苗的病害也较为严重。本研究通过对离体培养中基本培养基、激素、糖类、pH值、光照、附加物、外植体等因素的探索,优化了安祖花快速繁殖体系,提高了愈伤诱导率和不定芽分化率;对安祖花进行移栽前药剂浸泡和炼苗处理,提高了移栽成活率,探讨了栽培过程中基质、光照等因素的影响,提高了种苗的生长速度和质量;对安祖花栽培中出现的细菌性枯萎病的病原菌进行了初步鉴定,运用链霉素抑制了病害的发生。其研究结果如下:
1 安祖花大规模离体培养快速繁殖技术的优化
在安祖花的愈伤组织诱导过程中发现,将BA与2.4-D配合使用比单独使用BA或2.4-D效果要好,诱导率分别提高8%和30%左右。适当降低基本培养基中N元素的浓度可以提高愈伤诱导率,当NH_4NO_3与KNO_3降到250mg/L时,诱导率达到100%,比前人的研究提高15%左右。使用不同糖类,诱导率不同,使用30g/L蔗糖或葡萄糖诱导率均可达到100%,比使用麦芽糖高13%;而在愈伤质量上,用葡萄糖诱导的愈伤组织较大,质量较好。以无菌苗叶片、叶柄为外植体,不同品种诱导率均可达97%以上;而盆栽苗则以全展叶的叶柄和成熟叶的叶片为外植体最好。愈伤组织诱导培养基以改良MS+BA0.2(0.5,1.0)+2.4-D0.1+30g/L葡萄糖较好。
在愈伤组织分化不定芽时,使用较高浓度的BA与NAA,分化率可达到91.7%,而通过使用葡萄糖作为碳源,可使分化率提高到96.1%。分化培养基以改良MS+BA2.5+NAA0.5+30g/L葡萄糖较好,分化率高,分化芽较多。
以腋芽增殖途径繁殖试管苗,培养基以改良MS+B ALO王NAA 0.1较好,增殖倍
数达6.6,芽较长、基部愈伤少:
在安祖花试管苗继代增殖过程中,基本培养基对继代产生影响,在试管苗鲜重
的增长率上,改良MS与MS之间具有极显著差异。椰乳对生长具有促进作用,与
不含椰乳的培养基之间差异显著。强光有利于试管苗的生长,光照强度为5500L况
与2500Lx对试管苗鲜重增加率具有极显著差异。较适合的继代培养基为改良MS
+BA(KT)2.。+NAAO.05+葡萄糖3。叭+椰汁100叭。在此继代培养基上,试管
苗还能长出3一5条粗壮的根,可以不经生根直接进行移栽,简化了培养程序.
2提高安祖花试管苗移栽成活率及生长速度的研究
在试管苗移栽前的处理中,多菌灵溶液浸泡处理比KNln04溶液处理效果要好,
其中,多菌灵以0.2%处理20min较好,成活率达91 .1%,比KMno4溶液处理的要
高12%。
揭膜炼苗对试管苗适应外界环境具有好处,揭膜5天成活率达到86%,揭膜7
天以上,成活率达98%。移栽时带3条或3条以上的根对移栽苗生长时的株高、叶
片数、根数均具有促进作用。
从株高、叶片数、根数方面综合考虑,移栽基质以椰子壳:泥炭土:珍珠岩为
2:0.5:0.5为好。在株高、叶片数、叶面积、根数上,普通日光灯和植物生长灯之间
没有显著差异。
护
3新发现的安祖花细菌性枯萎病致病菌的初步鉴定与防治
从安祖花枯萎病病叶上分离和纯化,得到一株细菌,通过回接安祖花,发现此
细菌为安祖花细菌性枯萎病的致病菌。此细菌为革兰氏阴性菌,通过Biolog
Microstatinn细菌自动鉴定系统将此菌株鉴定为恶臭假单胞菌生物变种A
(Pseudomonas put才da biovarA)。此菌对安祖花的危害未见有报道,因此,此菌为一
种新发现的安祖花细菌性枯萎病的致病菌。
链霉素对此细菌的抑制作用比氯霉素要好,80mglL的链霉素即可完全抑制此细
菌。通过用农用链霉素(1000万单位,149)配制成的0.15%的溶液进行试管苗移栽
前的浸泡处理,最终成活率可以达到94.6%;对移栽苗进行灌根和叶面喷施,对病
害的抑制具有较好的效果。
Anthurium andraeanum, a kind of famous and precious flower introduced to China in recent years, has become the favorite of most people for its peculiar appearance, long flurescense and rich colors. However, the development ofAnthurium andraeanum sector in China was still far from booming due to the backward traditional method of propagation, which was limited to special seasons and of low propagation rates. Tissue culture, applied to rapid propagation of large scale in flower industry, was considered a possible way for Anthurium andraeanum. Unfortunately, many former researchers had tried hard but failed in it, especially in plantlets hardening, transplanting and disease controlling. After systematic researches on related factors such as explants, basic media, hormones, carbon resources, pH, light and additives, the traditional techniques of Large-scaled tissue culture of Anthurium andraeanum in promoting calli induction and buds differentiation were optimized successfully. Survival rate of transplanting was
improved through chemical soaking and adaptive training before transplanting, and growth speed was accelerated through discussing the influences of substance, illumination, etc. At last, the pathogenic bacterium of blight was identified initially and was inhibited by streptomycin. The results as flowing:
1. Improvement on tissue culture techniques of Anthurium andraeanum
For calli inducing, a combination of BA-6 with 2.4-D was more effective than single of them, enhancing inducing rates by about 8% and 30% respectively. The rate of calli inducing also rose up to 100% hi the improved medium with reduced N to 250mg/l in the form of NH4NO3 and KNO3, 15% more than that of previous researches. Different forms of hydrocarbon resources also led to different calli inducing rates and calli qualities. Calli inducing rate could reach 100% in 30g/l sucrose or glucose, 13% higher than in maltose, and the quality of induced calli hi glucose was much better than hi others. Taking leaves and petioles of sterile plantlets as explants, the inducement rates of different varieties were more than 97%. Taking materials from pot plantlets, the petioles of frondesces and blondes of ripe leaves were suitable to induce calli. Modified MS+ BA0.2(0.5orl.O) +2.4-D0.1 +glucose30g/l was proved to be the proper medium for calli induction.
Comparatively higher concentration of BA and low concentration of NAA had positive effects on buds differentiation, and it could be up to 96.1% by glucose as carbon resource. Modified MS+BA2.5+NAA0.5+glucose30g/l was the proper medium for buds differentiation, the quality of buds was better, differentiation rate was higher and quantity was more.
Axillary buds, another kind of ideal explant, multiplied by 5.6 times in modified
MS+BA1.0+NAA0.1.
Basic medium had effects on subculture. Rates of increase on fresh weights of test tube plantlets on modified MS and MS were different extremely remarkably. The culture media containing coconut milk promoted the growth far more effectively than non-coconut milk ones. Strong light was also a stimulus for the growth of test tube plantlets and in the light of 2500Lx to 5500Lx, the increase rates of fresh weight was much higher than those in weaker light. The modified MS+BA(KT)+NAA0.05 +glucose30g/l +CM100g/l was proved to be proper medium for subculture, in which the test tube plantlets were much stronger and 3-5 roots could be induced from it. This kind of plantlets could be transplanted directly.
2. Studies on factors to transplanting survival rate and growth
The Carbendazin-sulfar was superior to KMnO4 in dipping treatment of test tube plantlets against infection before transplantation. 20 minutes such treatment m 0.2% of Carbendazin-sulfar made the survival rate up to 91.1%, 12% higher than that of KMnO4.
Exposed to the outside environment for 5 days by uncovering plastic mantle test tube plantlets could survive by 86% and more than 7 days exposure led to 98% survival. The test tube plantlets with 3 or more roots often grew better. According to increment of
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