【作者】 王静华；

【导师】 刘桂林； 梁海永；

【作者基本信息】 河北农业大学 ， 园林植物与观赏园艺， 2009， 硕士

【摘要】 榆树是一种分布广泛的古老树种,也是著名的行道树和庭院绿化树种,在我国素有“杨柳榆槐”之说。随着对榆树研究的发展和深入,尤其是育种工作对遗传基础探索的迫切要求和分子生物学研究成果对传统遗传育种的影响和巨大冲击,国内外许多研究已经逐渐转向榆树的遗传学基础。建立一个成熟的组织培养及再生体系是进行遗传转化的先决条件。本研究以两个不同种源地榆树的种子为材料进行了组织培养,并建立了较为完善的再生体系,为榆树的生物技术改良和优良品系的快速繁殖提供了技术基础。主要研究结果如下:1.启动培养时以两种榆树的种子作为外植体,以MS培养基为启动培养基,最佳的灭菌方式是:20%的84消毒液灭菌15 min。2.以两种榆树的带芽茎段为材料研究了基本培养基、生长调节剂、培养基pH值和蔗糖浓度对增殖培养的影响,筛选出了最适宜的增殖培养基:河北沧州榆树为MS+6-BA 0.10 mg/L+IBA 0.005 mg/L,蔗糖20 g/L,pH值5.8~6.2;内蒙赤峰榆树为MS+6-BA 0.10 mg/L+IBA 0.025 mg/L,蔗糖30 g/L,pH值5.8~6.2。在增殖培养的多个处理中内蒙赤峰榆树的增殖系数高于河北沧州榆树。3.以两种榆树的叶片为材料,通过诱导叶片产生愈伤组织,再诱导愈伤组织分化的途径,得到了生长健壮的不定芽。筛选出了分别适宜两种榆树的生长调节剂组合。诱导榆树叶片产生愈伤组织的最适培养基:河北沧州榆树为MS+TDZ 0.025 mg/L+IAA 0.1 mg/L,出愈率为93.00%;内蒙赤峰榆树为MS+TDZ 0.05 mg/L+IAA 0.1 mg/L,出愈率为98.26%。两种榆树的愈伤组织在MS+TDZ 0.05 mg/L+IBA 0.025 mg/L的培养基上培养时,宜分化产生不定芽,且不定芽生长迅速。河北沧州榆树的再生频率为52.20%,平均出芽数4.02;内蒙赤峰榆树的再生频率为55.04%,平均出芽数3.87。4.以榆树叶片为材料,研究不同生长调节剂组合对诱导其直接分化产生不定芽的影响。结果表明,两种榆树诱导叶片直接产生不定芽的最佳培养基均为MS+TDZ 0.005 mg/L+IAA 0.01 mg/L。河北沧州榆树的再生频率为68.35%,平均出芽数为4.89;内蒙赤峰榆树的再生频率为72.59%,平均出芽数为4.91。5.两种榆树叶片直接分化的再生频率均高于通过愈伤组织诱导分化的再生频率。所以榆树最佳的再生途径是诱导叶片直接分化。6.以榆树再生苗茎段为材料,研究不同生长调节剂组合对生根培养影响,筛选出了榆树再生苗最适生根培养基:河北沧州榆树为MS+IBA 0.01 mg/L,内蒙赤峰榆树为MS+IBA 0.05 mg/L。在生根培养的多数处理中内蒙赤峰榆树不如河北沧州榆树易于生根,内蒙赤峰榆树的最高生根率为89%,也低于河北沧州榆树的100%。更多还原

【Abstract】 The Siberian elm, one kind of widespread distributed ancient species,the famous roadside and the garden afforestation tree, was known as“Poplar, Willow, Siberian elm & Sophora Japonica”in our country saying. Along with the development of Siberian elm research, particularly the urgent need during breeding work on the hereditary basis and the influence and huge impact of molecular biology research to the traditional heredity breeding, many domestic and foreign research already gradually changed into the genetics basic of Siberian elm. To establish a mature tissue culture and regenerates system is a precondition for gene transformation. An tissue culture research has been conducted through taking two different provenances of Siberian elm’s seeds as the primary material in this work and a fine regeneration system has been established which provide a technology support for Siberian elm’s biological technology improvement and the fine strain’s fast reproduction. The main findings are as follows:1. Two different provenance Siberian elm’s seeds had been used as primary explants and MS medium had been used as initiation medium during initiation culture. The best sterilization method is: 20%“84 Xiaoduye”15 minutes.2. The influence of basic medium, growth regulator, medium pH and sucrose concentration on regeneration has been studied by using stem with buds from two different provenances of Siberian elm. The best regeneration medium has been selected out as: MS+6-BA 0.10 mg/L+IBA 0.005 mg/L, sucrose 20 g/L and pH 5.8~6.2 for Siberian elm from Cangzhou Hebei; MS+6-BA 0.10 mg/L+IBA 0.025 mg/L, sucrose 30 g/L and pH 5.8~6.2 for Siberian elm from Chifeng Inner Mongolia. The regeneration coefficients of Siberian elm from Chifeng are higher than the elm from Cangzhou in many treatments.3. Using leaves from two different provenances Siberian elm as material, healthy adventitious buds were got through the process that callus inducing from leaves followed with differentiation inducing from callus. Combinations of growth regulator suitable for these two elms have been selected out. The optimum medium for inducing callus from leaves is: MS+TDZ 0.025 mg/L+IAA 0.1 mg/L for elm from Cangzhou with a callus formation rate 93.00%; MS+TDZ 0.05 mg/L+IAA 0.1 mg/L for elm from Chifeng with a callus formation rate 98.26%. It was easy to get adventitious bud followed with a fast bud growth for callus from these two kinds of elm when cultured at the medium MS+TDZ 0.05 mg/L+IBA 0.025 mg/L. The regeneration rate is 52.20% for elm from Cangzhou and 55.04% for elm from Chifeng; the average number for buds formation is 4.02 for elm from Cangzhou and 3.87 for elm from Chifeng. 4. Using leaves from Siberian elm as material, the effect of different combinations of growth regulator on inducing adventitious bud directly from leaves has been studied. The results indicated that the optimum medium for this work in these two Siberian elms is MS+TDZ 0.005 mg/L+IAA 0.01 mg/L. The regeneration rate is 68.35% and the average bud formation is 4.89 for elm from Cangzhou; the regeneration rate is 72.59% and the average bud formation is 4.91 for elm from Chifeng.5. For both of the two different provenances Siberian elm, the regeneration rates are higher for the process through direct differentiation from leaves than the process through callus inducing differentiation, therefore, the optional regeneration approaches for Siberian elm is to induce differentiation from leaves directly.6. Using regenerated stem from Siberian elm as material, effect of different combinations of growth regulator on rooting has been conducted and the optimum rooting medium has been selected out: MS+IBA 0.01 mg/L for elm from Cangzhou; MS +IBA 0.05 mg/L for elm from Chifeng. Among all these rooting processes, in most case, it was hard to form root for elm from Chifeng than from Cangzhou and 89%, the highest rooting rate for elm from Chifeng, was also lower than the 100% for elm from Cangzhou.更多还原