【作者】 张宗勤；

【导师】 王跃进；

【作者基本信息】 西北农林科技大学 ， 果树学， 2004， 硕士

【摘要】 采用不同培养基、不同种类、不同浓度的细胞分裂素与生长素组合，对19个葡萄品种或株系的叶片、叶柄、茎段及根段进行了组织培养研究，取得的主要结果如下： 1、葡萄外植体无菌系建立：去除顶芽后的葡萄新梢第1～3节为适宜的外植体材料。在不适宜取材的夏秋冬季，采用二次间歇式消毒能有效抑制外植体的污染，降低褐变率。2、胚性愈伤组织与胚状体的诱导：诱导供试多数葡萄离体叶片、茎段、根段外植体脱分化的适宜培养基是NN69+2,4-D 2.0 mg/L+BA 0.2 mg/L，叶柄以NN69+2,4-D 2.0 mg/L + BA 2.0 mg/L为宜。试管苗顶部1～3片叶再生容易，幼叶在多种培养基上能全部愈伤化；而成熟叶片仅自叶脉割伤处与叶柄切口处产生愈伤组织。茎段在切口部位首先产生愈伤组织，茎表皮组织先局部，进而整个茎段愈伤化。供试品种中，无核白、红宝石无核、红光无核、底来特、爱莫无核、无核紫、红无籽露、红葡萄、早玫瑰、黑奥林、红地球、左山1号共12个品种或株系的叶片或叶柄外植体经脱分化形成了胚性愈伤组织及胚状体。多数葡萄品种或株系体细胞胚胎发生的适宜培养基为1/2 B5+BA 5.0 mg/L + IBA 0.1 mg/L。NN69+2,4-D 0.1 mg/L +BA 0.2 mg/L适宜于胚性愈伤组织的保持培养。1/2 B5+BA 0.2 mg/L +CH 100 mg/L 的培养基适宜于胚状体萌发。3、不定芽的诱导：经不定芽途径再生植株的品种分别是无核白、红光无核、红宝石无核、红葡萄、杨格尔、底来特、无核紫、红地球、红无籽露和力扎马特。基因型不同，叶片外植体不定芽再生的适宜培养基不同，如无核白、红宝石无核为1/2 MS+BA 5.0 mg/L +TDZ 0.1 mg/L +IBA 0.05 mg/L，红葡萄为NN69+BA 5.0 mg/L +TDZ 0.2 mg/L，红光无核为1/2 MS+BA 1.0 mg/L +TDZ 0.02 mg/L；无核紫为NN69+BA 1.0 mg/L +TDZ 0.1 mg/L +IBA 0.1 mg/L。同一品种或株系的外植体不同，不定芽再生的适宜培养基也不同，如红宝石无核叶柄再生的适宜培养基为1/2 MS+BA 5.0 mg/L +TDZ 0.1 mg/L +IBA 0.1 mg/L。红宝石无核不定芽发生率最高（36.67%），平均每个外植体产生2.27个不定芽。4、再生植株的生根培养：葡萄不定芽生根的适宜培养基为1/2 MS+IBA 0.1 mg/L。在较强光照条件下（3000 Lx），经1个月驯化培养可使植株健壮，达到移栽条件。更多还原

【Abstract】 Experiments of initiation and regeneration of varieties of grapevines in vitro were performed. Ten and 12 of all 19 varieties involved in this study were obtained adventitious buds and/or embroids respectively through distinguished liquid or agar solid media combined with some kinds of, concentrations and combinations of plant growth regulators. Effective regeneration experimental technique series of different types of grapevine varieties were established via studying of factors affecting organogenensis and embryogenensis, for meeting further genetic transformation operations. 1. Initiation of grapevines: Sample season was the critical factor influencing the initiation efficiency. The lowest contamination was obtained at spring, April to May, and the highest was at august. The suitable explants were the top 1~3 node sections of plants (apex excised) compared with the 4~6 nodal parts. Dedicate terminals were not explants needed because of their higher browning probability. Twice sterilizing technique with 48h interval wan the top efficiency of initiation of 90%, which was 30-40 percent higher than regular one step sterilization sequence. 2. Embryogenesis: The appropriate medium for dedifferentiation was NN69+2,4-D 2.0 mg/L + BA 0.2 mg/L for leaf, shoot and root segment explants, was NN69+2,4-D 2.0 mg/L +BA 2.0 mg/L for petiole cut. Embryogenic callus were initiated on NN69+2,4-D 2.0 mg/L +BA 0.2 mg/L of leaf explants. Embroids were obtained on 1/2 B5+BA 5.0 mg/L + IBA 0.1 mg/L, of Thompson Seedless, Flame Seedless, Delight, Olmo Seedless, and Zuoshan-1. Embryogenic callus could be maintained on NN69+2,4-D 0.1 mg/L +BA 0.2 mg/L. The appropriate medium for embroid maturation was 1/2 B5+BA 0.2 mg/L +CH 100 mg/L.3. Organogenesis: Adventitious buds were obtained of 10 of 19 varieties involved, Thompson Seedless, Flame Seedless, Delight, Youngle, Red Globe et al. The optimal medium for leaf disc explants organogenensis for ‘Thompson Seedless’ and ‘Ruby Seedless’ were 1/2 MS+BA 5.0 mg/L +TDZ 0.1 mg/L +IBA 0.05 mg/L, for ‘Red Grape’ was NN69+BA 5.0 mg/L +TDZ 0.2 mg/L，for ‘Flame Seedless’ was 1/2 MS+BA 1.0 mg/L +TDZ 0.02 mg/L. The highest regeneration ratio was obtained at 36.67% of ‘Ruby seedless’ leaf explants.4. Rooting: Rooted plants could be obtained of plantlets regenerated, of 1.5~2 cm high or more, at 1/2 MS+IBA 0.1 mg/L. They were ready to be transplanted after 1 month 3000 Lx lighted acclimation.更多还原