【作者】 王刚；

【导师】 幸亨泰；

【作者基本信息】 西北师范大学 ， 植物学， 2002， 硕士

【摘要】 百合（LiLium spp）是单子叶植物亚纲百合科（LiLiaceae）百合属（Lilium）的所有种类的总称。百合的鳞茎含有丰富的营养，具有很高的食用价值；中医普遍认为百合入药具有补中益气、养阴润肺、止咳平喘的功效；百合也是一种名贵切花，其花形美丽、花色多样，是继鹤望兰和红掌之后的又一只切花新秀。百合的快繁主要有小鳞茎分株繁殖、鳞片扦插及组织培养。本实验利用百合属的兰州百合（Liliun davidii var.Unicolor（hong）cotton）及野百合（Lilium brownii F.E. brown ex Miellez）进行组织培养，从外植体的取材、消毒、初代培养物的建立、激素的种类及浓度的筛选、芽的增殖、根的诱导、移栽及组织培养过程中两种百合染色体的行为和B染色体进行了研究，初步提供了一组用于两种百合组织培养的培养基，阐述了组织培养过程中两种百合染色体的变化和B染色体的有无。 在两种百合的初代培养时，外植体在2001年春季取回。鳞片取回后用毛刷刷去泥土，流水冲洗24h。材料接种前在超净台上用75%酒精浸泡30s后，于0.1%升汞溶液中消毒13min，最后用无菌水冲洗5-6次。将消完毒的兰州百合及野百合外植体切成0.5cm*0.5cm的方块,分别接种于MS+0.6-1.0mg/lBA+0.2mg/LNAA及MS+0.1-0.5mg/lBA+0.1-0.2mg/LNAAH或0.5mg/lIAA的诱导培养基上，诱导率可高达70-100%，并发现外植体有以外层下部与鳞茎盘相连的部分为最佳外植体。增殖培养时，兰州百合的最佳增殖培养基为MS+0.5mg/lBA+0.1mg/lNAA，而MS+0.6mg/lBA+0.1mg/lNAA及MS+1.0mg/lBA+0.1mg/lIAA也有较好的诱导效果，野百合的最佳增殖培养基为MS+1.0mg/lBA+0.1mg/LIAA,而MS+0.7mg/LBA+0.2mg/lNAA及MS+0.9mg/lBA+0.3mg/lNAA也较理想。在23-25℃，2000LX，16h光照，8h黑暗或12h黑暗，12h光照条件下培养，每28-32d左右继代一次，继代6次左右，增殖倍数可达4-5倍。 生根培养时，以1/2MS为基本培养基，附加0.1-0.2mg/lNAA或IAA可使根长的长而壮，活性炭对根的生长有利。根长至1-1.5cm时，开瓶炼苗3-5d后，移栽至盛有蛭石加腐殖土（1:1）的纸杯中，保持70%左右的湿度，60%的自然光，成活率可达80%以上，且移栽后长势良好。 组织培养时染色体存在一定的变异。愈伤组织中兰州百合及野百合的变异率分别为54.4%和49.4%，染色体数目变异范围分别为9-50和12-50条；而芽端分生组<WP=5>织中兰州百合及野百合的变异率分别为17.5%和9.5%，说明愈伤组织中染色体的变异率高于芽端分生组织。激素对组织培养过程中染色体行为有明显的影响，单独使用一种激素如NAA引起的兰州百合及野百合染色体的变异率为3.5%及2.8%，而BA和NAA搭配引起的变异率普遍高于此值。另外不同种类及浓度的激素搭配引起的变异率也不相同。综合诱导率及变异率两种因素认为，No.5及No.11分别为兰州百合及野百合的最佳诱导培养基。组织培养时，继代次数不宜过多。实验中发现，继代次数增多导致染色体变异率的增加，染色体变异率增加使芽的增殖能力降低。兰州百合继代以4-5次较好，野百合以6次为好。不同基因型的百合在组织培养过程中染色体变异率无显著差异。根尖细胞中染色体变异率显著低于组织培养过程中各种组织的细胞。 对兰州百合及野百合的根尖细胞、愈伤组织及芽端分生组织细胞进行普通压片观察，发现野百合各种组织细胞中无B染色体被观察到；兰州百合的根尖、愈伤组织及芽端分生组织细胞中有B染色体，数目为1-2条。更多还原

【Abstract】 This article research the tissue culture and cloning propagation and the chromosomal variation of two kinds of lilies —Lilium davidii Var.Unicolor(Hoog)Cotton and Lilium brownii F.E.Brown ex miellez.At the same time, we also observed the nuclear and the B-chromosome. The tissue culture and cloning propagation include the establishment of aseptic culture, bud induction and proliferation, selection of suitable hormone , rooting and acclimatization. The chromosomal variation include the change of chromosome number mainly.When the explants were collected in the spring, high induction rate can be achieved. Brushing the dust and washing the explants under ranning tap water to remove some of the dirt and contaminations, in 75%CH3CH2OH for 30 second and in 0.1%Hgcl2 for 13 minutes, rinsed 5-6 times with sterile water. The two kinds of explants were cultured on Murashige&Skoog medium(MS) added with 0.6-1.0mg/lN-benzyladenine(BA)+0.2mg/l naphthalane acetic(NAA)and added with 0.1-0.5mg/lBA+0.1-0.2mg/lNAA or indole-3-acetic(IAA).The induction rate can reached 70-100%.The best explants were the parts that were below and linked the scales.During the buds poliferation,the best medium of L.davidii Var.Unicolor(Hoog)Cottonis MS+0.5mg/lBA+0.1mg/lNAA, the best medium of L.brownii F.E.Brown ex miellez is MS+1.0mg/lBA+0.1mg/lIAA. It good to keep the culture at 23-25℃,photoperiod with a light intensity of 2,000Lx,keep 16 hours lighting and 8 hours darking or 12 hours lighting and 12hours darking on culture, to subculture every 28-32days.The poliferation can be reached 4-5 times.One-half MS basal medium added 0.1-0.2mg/lNAA or IAA can make the root long and strong. The activated charcoal is good to root formation and growth. When the length of root is 1-1.5cm,lilies can be acclimatized. If the living condition is good, the survival rate were over 80%During the tissue culture, we found the chromosomal variation in cells of two kinds of lilies. The variation rate of callus of L.davidii Var.Unicolor(Hoog)Cotton is 54.4%and the L.brownii F.E.Brown ex miellez is49.4%, the chromosome number varied from 9-50<WP=7>and 12-50;the variation rate of buds of L.davidii Var.Unicolor(Hoog)Cotton and L.brownii F.E.Brown ex miellez are 17.5%and 19.5%,these show that the variation rate of callus is higher than buds. The hormone can affect the deeds of the chromosome. The variation rate caused by single hormone is lower than two kinds of hormone, the kind and intensity of hormone is different, the variation rate is different. We think that No.5and No.11 are best medium to two kinds of lilies when we thought over the induction rate and variation rate. The times of subculture should be limited because we found when the subculture times is increasing, the variation rate is higher and the differentiation capacity is lower. The best times of L.davidii Var.Unicolor(Hoog)Cotton is 4-5,the other is 6 times.We observed the cells of root-tip, callus and bud of two kinds of lilies, we could not find the B-chromosome in cells of L. brownii F.E. Brown ex miellez, but find it in L.davidii Var.Unicolor(Hoog)Cotton,the number is 1-2.更多还原