【作者】 汤访评；

【导师】 陈发棣；

【作者基本信息】 南京农业大学 ， 观赏园艺， 2009， 博士

【摘要】 菊花具有很高的观赏和经济价值,被广泛用作切花、盆栽和公园花坛布置。但许多菊花品种对病害、虫害和其它环境胁迫缺乏抗性。菊花近缘属中存在许多如抗病、抗虫、抗逆等优良特性的优异种质。本研究首先以菊属(Dendranthema)大岛野路菊(D.crassum(Kitam.)Kitam:2n=10x=90).南京野菊(D.indicum(L.)Des Moul; 2n=4x=36)和菊花脑(D.nakingense(Hand.-Mazz.) Y.R.Ling;2n=2x=18)为母本,分别与亚菊属(Ajania)多花亚菊(A.myriantha(Franchet) Ling ex Shm; 2n=2x=18)、芙蓉菊属(Crossosstephium)芙蓉菊(C.chinense(L.)Makino;2n =2x=18)、太行菊属(Opisthopappus)太行菊(O.taihangensis(Ling)Shih;2n= 2x=18)和菊蒿属(Tanacetum)菊蒿(T.vulgare L.;2n=2x=18)进行属间远缘杂交,探讨菊属与近缘属植物间亲缘关系；再通过属间杂种F1为桥梁亲本与栽培菊花杂交,进行菊花种质创新,为抗性菊花新品种选育奠定基础,主要研究结果如下：(1)以大岛野路菊和南京野菊为母本,芙蓉菊为父本进行属间杂交没有得到种子。大岛野路菊和南京野菊分别有57%和46%的大孢子母细胞能发育成成熟的雌配子体。父本芙蓉菊花粉离体萌发率为51%,授在母本柱头上2h开始萌发,一直延续2 d,说明花粉活力较高。通过对授粉后不同时间段子房切片的观察,发现子房中几乎没有胚胎,说明芙蓉菊与大岛野路菊和南京野菊远缘杂交失败的主要原因可能是由杂种胚的早期败育所致。(2)对12个属间杂交组合授粉后15d的大量胚珠进行离体培养,共获得大岛野路菊×芙蓉菊、南京野菊×芙蓉菊、大岛野路菊×多花亚菊、菊花脑×菊蒿及南京野菊×太行菊5个杂交组合后代,其中前3个组合各分别获得1个株系,后两个组合分别获得4个和7个株系。而在常规人工杂交条件下,这5个杂交组合均未获得杂种后代,表明胚珠拯救是获得菊属与近缘属远缘杂种的一条有效途径。(3)对5个属间杂种后代进行基因组荧光原位杂交分析,发现菊属与亚菊属基因组在荧光原位杂交时需要高浓度的封堵DNA才能区分彼此,说明菊属与亚菊属之间亲缘关系较近；而菊属与芙蓉菊属、菊蒿属和太行菊属的基因组在荧光原位杂交时不需要封堵DNA,说明彼此间亲缘关系相对较远。此外,(南京野菊×芙蓉菊)F1生长健壮,开花茂盛；(南京野菊×太行菊)F1生长健壮,但不见花芽分化；(南京野菊×菊蒿)F1在田间无法正常生长。推测菊属与4个近缘属间的亲缘关系由近及远依次为亚菊属、芙蓉菊属、太行菊属和菊蒿属。(4)以栽培菊花品种‘天坠玉露’为母本,大岛野路菊×芙蓉菊、南京野菊×芙蓉菊和菊花脑×菊蒿3个属间杂种F1为父本进行人工杂交,发现结实率分别为22.5%、12.5%和0.25%。对杂种后代进行基因组双色荧光原位杂交分析,在‘天坠玉露’×(南京野菊×芙蓉菊)F1和‘天坠玉露’×(菊花脑×菊蒿)F1杂交后代中没有观察到芙蓉菊和菊蒿染色体,说明两个近缘属植物的染色体可能已被排斥,但在‘天坠玉露’×(大岛野路菊×芙蓉菊)F1杂交后代中观察到了‘天坠玉露’、大岛野路菊和芙蓉菊3个基因组染色体,实现了栽培菊花种质创新,使充分利用菊花近缘属优良种质进行栽培菊花品种改良成为可能。更多还原

【Abstract】 Chrysanthemum(Dendranthema morifolium), is one of the most important floricultural crops in cut flower, potted plant and garden plant. Many chrysanthemum cultivars are weak resistant to disease, insect, virus and environment. But in allied genera, many wild species possess elite attributes such as resistance to biotic or abiotic stresses. In this paper, intergeneric cross has been made to assess the relationships between Dendranthema wild species (♀) and C. chinense (2n=18;(?)), A. myriantha (2n=18;(?)), T. vulgare (2n=18;(?)), O. taihangensis (2n=18;(?)) respectively. Then, as bridge parent, the intergeneric hybrids were hybrided with chrysanthemum to innovate chrysanthemum germ plasm. It made a base for new resistance varieties breeding of chrysanthemum and the results showed as below:(1) No seed was obtained in the combination between C. chinense((?)) and D. indicum (♀), D. crassum (♀).57% and 46% megaspores of D. indicum and D. crassum can develop into a mature embryo sac. In vitro,51% C. chinense pollens germinate, and in planta, germination is observed from 2h to 2d after pollination. And, the development of the hybrid embryo was only arrested at the globular stage. It seems that hybrid embryo abortion is the major means of preventing the formation of intergeneric hybrid combination.(2) Aapplying of ovule rescue with 12 combinations after pollination for 15d, the combination of D. crassum and C. chinense, D. indicum and C. chinense, D. crassum and A myriantha, D. indicum and O. taihangensis and D. nankingense and T. vulgare were obtained hybrid plants. Under natural conditions, no seed was obtained in these combinations. Results showed that ovule culture is a valid method for carrying out and overcoming distant hybridization between Dendranthema and the allied genera.(3) Genomic in situ hybridization experiments were able to successfully distinguish the genomic origin of both mitotic and meiotic metaphase chromosomes in the hybrid. In D. crassum×A. myriantha, a large excess of blocking DNA is necessary to achieve genome discrimination in the hybrid. In contrast, in D. crassum×C. chinense, D. indicum×C. chinense, D. indicum×O. taihangensis, and D. nankingense×T. vulgare hybrids, no blocking DNA is needed to fully discriminate between the parental genomes. GISH analysis suggested distant phylogenetic relationship between Dendranthema and Crossostephium, Tanacetum, Opisthopappus, while Dendranthema is more close to Ajania than the others.The D. indicum×C. chinense hybrid grew and abloomed vigorously in the field, while the D. indicum×O. taihangensis hybrid could not carried out genital growth and D. indicum×T. vulgare hybrid could not grow normally. It showed that the phylogenetic relationship between Dendranthema and the allied genera from close to distant in turn is Ajania, Crossostephium, Opisthopappus, Tanacetum.(4) Artificial hybridizations have been made between the Chrysanthemum cultivar ’tianzhuiyulu’(2n=54;♀) and the intergeneric hybrids ((?)). The seed set of ’tianzhuiyulu’×(D. Crassum×C. chinense),’tianzhuiyulu’×(D. indicum x C. chinense) and ’tianzhuiyulu’×(D. nankingense×T. vulgare) was 22.5%,12.5% and 0.25% respectively. Dual-color genomic in situ hybridization was applied for these hybrids. In ’tianzhuiyulu’×(D. indicum×C. chinense) hybrid, it contained two genomes of ’tianzhuiyulu’ and D. indicum. In ’tianzhuiyulu’×(D. nankingense×T. vulgare) hybrid, it contained two genomes of ’tianzhuiyulu’ and D. nankingense. While in ’tianzhuiyulu’×(D. crassum×C. chinense) hybrids, they contained three genomes of ’tianzhuiyulu’, D. crassum and C. chinense, the artificial hybridizations was an effective way to innovate chrysanthemum germplasm and made it possible for improving chrysanthemum varieties by introducing elite attributes of its allied genera.更多还原