卫矛组织培养技术研究

发布时间：2018-04-23 15:18

  本文选题：卫矛 + 不定芽　； 参考：《沈阳农业大学》2017年硕士论文

【摘要】：本研究以卫矛(Euonymus alatus(Thunb.)Sieb)不同部位为外植体,探讨了取材时间及灭菌方法对茎段消毒效果的影响;研究了沙藏与未沙藏、成熟与未成熟、带假种皮与去除假种皮及GA3浓度4种因素对种子萌发的影响,沙藏与未沙藏的成熟种胚、NAA(0.5、1.0、2.0mg/L)和 6-BA(0、1.0mg/L)对不定芽诱导的影响,以及 NAA(0.5、0.05mg/L)、6-BA(1.0、2.0mg/L)和琼脂(6.5、7.0g/L)对茎段不定芽诱导的影响,探讨了基本培养基、2,4-D、TDZ、暗培养时间4个因子对花生殖器官(花萼、花瓣、雄蕊、雌蕊)和叶片的愈伤组织诱导的影响,从而筛选出最佳外植体及诱导方式;继而在基本培养基、6-BA、NAA、GA34个因子3个水平中选出不定芽增殖的最佳培养基;最终从添加NAA、IAA、IBA(0.2-4.0mg/L)及GA3(0.2-0.8mg/L)的28个处理中筛选出试管内生根最佳方法,并与试管外生根下的基质、生根剂、激素浓度的4个水平中的最佳生根方法进行了比较,筛选出卫矛最佳生根法。研究结果如下:(1)茎段最佳取材时间是5月中旬,染菌率为3%,存活率76.67%;最佳消毒方法是Hgc12 8min并加入两滴吐温-20,存活率59.67%。(2)去假种皮的沙藏种子在MS+GA32.0mg/L培养基中培养20d后萌发出子叶,种子萌发诱导率(10.2%)高于其他处理。含叶中脉的叶片在WPM+2,4-D 0.8 mg/l+ TDZ 1.0 mg/1培养基中的愈伤组织诱导率最高(65%-67%),但没有 B5+2,4-D 1.2 mg/l+ TDZ 2.0 mg/l+暗培养 20d处理中的雌蕊愈伤组织诱导率(均值86.5%)高。雌蕊愈伤组织为浅绿疏松状,并不是最佳愈伤组织形态。因而用卫矛叶片、雌蕊作为外植体诱导愈伤组织未取得成功,还有待研究。种胚不定芽诱导的最佳培养基为MS+6-BA1.0mg/L +NAA0.5mg/L,不定芽诱导率为12.8%。而茎段在6-BA2.0mg/L+NAA0.05mg/L培养基中的不定芽诱导率可达70.67%,远远高于种胚,因而茎段可为卫矛不定芽诱导的最佳外植体。(3)MS+6-BA1.0mg/L+NAA0.05mg/L是卫矛不定芽增殖的最佳培养基,增殖系数为23.51,即获得的丛生芽芽数为23.51个。在其上继代培养3次后,丛生芽芽数可增殖到48.5个。(4)卫矛试管内生根最佳培养基为1/2MS+GA30.4mg/L,生根率72.5%,平均生根数4.1条。试管外生根的最佳条件是蛭石+IAA2500mg/L,生根率为100%,平均生根数为5.3条。两种生根方法均可有效促使卫矛组培苗生根,但以试管外生根更佳。
[Abstract]:In this study, the different parts of Euonymus alatusus Thunb. Sieb were used as explants to investigate the effects of sampling time and sterilization methods on the sterilizing effect of stem segments, and to study the effects of sand storage and non-sand storage, mature and immature. The effects of four factors on seed germination, such as seed coat removal and seed coat removal and GA3 concentration, the effects of sand storage and non-sand storage on adventitious bud induction, and the effects of sand storage on adventitious bud induction, and the effects of sand storage and non-sand storage on adventitious bud induction, and the effects of NAA 0.55 mg / L 6-BA1.02.0 mg / L) and Agar 6.5g / L on adventitious bud induction. The effects of four factors on the callus induction of floral reproductive organs (calyx, petals, stamens, pistil) and leaves were studied. Then, the best medium for adventitious bud proliferation was selected from 3 levels of 34 factors of basic medium 6-BAANAA, and the best method of rooting in vitro was screened out of 28 treatments supplemented with NAA AIAA IBA 0.2-4.0 mg / L and GA _ 3N _ 0.2-0.8 mg / L), and the best rooting medium, rooting agent, was found with the substrate under rooting outside the tube, rooting agent. The best rooting method was screened out from the four levels of hormone concentration. The results were as follows: (1) the best sampling time of stem segment was in mid-May, the rate of bacterial infection was 3 and the survival rate was 76.67.The best disinfection method was Hgc12 8min and two drops of Tween -20, the survival rate was 59.67.2.) the seeds of sand-covered seed were cultured in MS GA32.0mg/L medium for 20 days and germinated to produce cotyledons. Seed germination induction rate was higher than other treatments. The callus induction rate of leaves with midvein was higher than that of B5224-D 1.2 mg/l TDZ 2.0 mg/l in WPM 24-D 0.8 mg/l TDZ 1.0 mg/1 medium, but the callus induction rate (mean 86.5) was higher than that of B5224-D 1.2 mg/l TDZ 2.0 mg/l dark culture for 20 days. The pistil callus is light green and loose, and is not the best callus morphology. Therefore, the callus induction with the leaves and pistil as explants has not been successfully induced, which remains to be studied. The best medium for adventitious bud induction was MS 6-BA1.0mg/L NAA 0.5 mg / L, and the rate of adventitious bud induction was 12. 8%. The adventitious bud induction rate of stem segment in 6-BA2.0mg/L NAA0.05mg/L medium was 70.67, which was much higher than that of seed embryo. Therefore, stem segment could be the best explant for adventitious bud induction of Leptanthus mongolicus L. 6-BA1.0mg/L NAA0.05mg/L was the best medium for adventitious bud proliferation. The multiplication coefficient was 23.51, that is, the number of buds was 23.51. After 3 times of subculture, the number of buds could proliferate to 48.5%. The best medium for rooting in vitro was 1/2MS GA30.4mg / L, the rooting rate was 72.5% and the average number of rooting was 4.1. The optimum conditions for rooting in vitro were vermiculite IAA 2500 mg / L, rooting rate was 100 and the average number of rooting was 5.3. The two rooting methods can effectively promote the rooting of the plantlets in vitro, but it is better to take root in vitro.
【学位授予单位】：沈阳农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S687
，

本文编号：1792515