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以花灌木华北绣线菊(Spiraea fritschiana)幼嫩健康的离体叶片为外植体,进行愈伤组织诱导、不定芽诱导及生根培养;再以华北绣线菊愈伤组织为受体材料,筛选选择抗生素和抑菌抗生素浓度,探讨菌液稀释倍数和侵染时间对根癌农杆菌遗传转化华北绣线菊的影响。结果表明,愈伤组织最佳诱导培养基为:MS+1.0 mg·L~(-1) 6-苄基腺嘌呤(6-BA)+0.1 mg·L~(-1)萘乙酸(NAA)+1.0 mg·L~(-1) 2,4-二氯苯氧乙酸(2,4-D),诱导率为94.16%;愈伤组织最佳增殖培养基为:MS+1.0 mg·L~(-1) 6-BA+0.03mg·L~(-1) NAA;不定芽最佳分化培养基为:MS+2 mg·L~(-1) thidiazuron(TDZ)+0.1 mg·L~(-1) NAA,分化率为96.11%;生根壮苗最佳培养基为:1/2MS+0.4 mg·L~(-1) NAA;以60 mg·L~(-1)卡那霉素(kan)为选择压、150 mg·L~(-1)头孢霉素(cef)为抑菌浓度,菌体用液体MS稀释2倍(OD_(600)=0.7),侵染8 min,为最优遗传转化体系,诱导抗性芽8.7个,遗传转化率为2%。本研究建立的华北绣线菊再生与遗传转化体系为其转基因研究及分子育种提供参考。
The young and healthy leaves of Spiraea fritschiana were used as explants for callus induction, adventitious bud induction and rooting culture. Then the calli were cultured on the callus of Spiraea fritschiana, The antibacterial and antibacterial antibiotics were screened and the effect of Agrobacterium tumefaciens genetic transformation on Spiraea polytricha was explored. The results showed that the optimal medium for callus induction was MS + 1.0 mg · L -1 6-BA +0.1 mg · L -1 NAA, 1.0 mg · L -1 2,4-dichlorophenoxyacetic acid (2,4-D), the induction rate was 94.16%. The best proliferation medium for callus was MS + 1.0 mg · L ~ (-1) 6-BA + 0.03 mg · L -1 NAA. The optimal medium for adventitious bud differentiation was MS + 2 mg · L -1 TDI +0.1 mg · L ~ (-1) -1) NAA, the rate of differentiation was 96.11%. The best medium for rooting was: 1 / 2MS + 0.4 mg · L -1 NAA; with 60 mg · L -1 kanamycin kan) was chosen as the antibacterial concentration of cefotaxime 150 mg · L -1 (cef). The bacterial cells were diluted twice with liquid MS (OD 600 = 0.7) and infected for 8 min Excellent genetic transformation system, induction of resistant buds 8.7, the genetic conversion rate of 2%. The regenerated and genetic transformation system of Spiraea polyrhina prepared by this study provides a reference for its transgenic research and molecular breeding.