Research On Techniques Of Lily (Lilium) Interspecific Hybridization And Resistant Breeding To Fusarium Wilt Disease

Lily (Lilium, 2n=2x=24) is not only an important cut-flower, pot plant and garden herb, but also used as a resource of food and medicine. Although China is the main distribution centre of lily species, the breeding work started recently. Hence, absorbing and introducing advanced technology and experience of lily breeding from abroad, breeding our own cultivars become increasingly necessary. Currently, the fast development of lily industry makes the cultivars from traditional hybrid groups can not meet the demand of consumers. As a result, breeding through interspecific hybridization is already one of the main aims in lily breeding.The wilting disease caused by Fusarium oxysporum f.sp. lilii is the most terrible disease in lily cut-flower and bulb production, resistant breeding to it is only sporadic reported in Asaitic hybrids. Since the fast development of lily industry, Asiatic hybrids can not meet the cunsumers demand any longer. As a result, Fusarium resistant breeding with Asiatic hybrids as crossing material is one of the most important breeding aims in lily. In addition, interspecific hybridization becomes the main way in current lily breeding. Hence, Fusarium resistant breeding combining with interspecific hybridization appears to be a completely new direction in lily breeding. This research, with the aim of guiding practical work, tries to explore new techniques for obtaining interspecific hybrids, disease test of Fusarium resistance & analysis of fertility in the obtained hybrids and method for meiotic polyploidization.In this thesis, the traditional cultivar from different hybrid groups, viz.'White Fox'from Longiflorum,'Conneticut King'from Asiatic,'Sorbonne'from Oriental and'Royal Goden'from Trumpet were selected, cut-style pollination and young embryo culturing methods were used for overcoming crossing barriers, followed by identification of the hybrids using molecular cytogenetic method, then fertility was investigated in the F1 hybrids of LA lily and pollen meiosis of 13 genotypes with partial fertility were observed, intergenomic recombination in backcrossing progeny were monitored and compared with the mitotic originated population using genomic in situ hybridization. Results showed: First, cut-style pollination significantly increased the crossing efficiency except the LL×TT combination, and the fruiting efficiency reached 65% in combiantion of LL×AA; the efficiency of embryo, embryo sac and ovule culture were 100%, 22% and 1% respectively. Results of genomic in situ hybridization (GISH) indicated them to be the real hybrids. It can be concluded that the cross results depends on the combination selected, embryo, embryo sac & ovule culture are efficient methods for hybrids obtaining, without false hybrid mixture. Embryo culture, although with a efficiency lower than embryo culture, is the predominant way of hybrids obtaining.Second, fertility was investigated in the F1 hybrids of LA lily with conventional cytological methods and thirteen genotypes of F1 hybrids of LA lilies (2n=2x=24) which showed partial fertility were used to illuminate the meiosis behaviors of homoeologous genomes using GISH. Results showed that a varied fertility was confirmed in 13 genotypes of LA hybrids; in these 13 genotypes with partial fertility, although the dissociation and abnormal segregation between homoeologous chromosomes leaded to the low fertility, variable number of bivalents existed within and between genotypes, and 12 bivalents in a few pollen mother cells (PMCs) could be detected in the genotypes that producing haploid gametes; diverse crossover events between non-sister chromatids existed in different PMCs; viable unreduced (2n), aneuploid and sporadic haploid (n) gametes were produced by these F1 hybrids of LA lilies; paracentric inversion heterozygote were also identified. It can be concluded that backcross the hybrids with their crossing parents will lead to triploid, aneuploid and diploid progeny.Third, two populations of Lilium hybrids viz. meiotic and mitotic polyploidized polyploids, were monitored and compared through cytological techniques. One of the populations was obtained through crossing an allotriploid Longiflorum×Oriental hybrid (LLO) and an allotetraploid Longiflorum×Trumpet hybrid (LLTT) both of which were derived from somatic chromosome doubling. The other type of allopolyploid population was derived from meiotic chromosome doubling in which numerically unreduced (2n) gametes from two different interspecific hybrids, viz., Longiflorum×Asiatic (LA) and Oriental×Asiatic (OA), were used to get backcross (BC) progeny with the Asiatic parents. GISH clearly discriminated the three constituent genomes (L, T and O) in the complements of the progeny obtained from mitotic chromosome doubling. A total of 26 genotypes were analyzed from this population and there was no evidence for any chromosomal rearrangements. However, in the case of meiotically doubled allopolyploid progeny considerable frequencies of chromosomal rearrangements were observed through GISH. The so-called chromosomal rearrangements in meiotic polyploids are the result of homoeologous recombination rather than"translocations". Evidence for the occurrence of meiotic recombination in the LA hybrids has been confirmed with GISH on meiotic chromosomes. Thus, there was evidence that neopolyploids of Lilium hybrids did not possess any noticeable chromosome rearrangements.Fourth, 99 interspecific F1 hybrids between Longiflorum and Asiatic hybrids were first evaluated for the Fusarium wilt disease resistance. Results indicated that the artificial infection of Fusarium oxysporum f.sp. lilii in greenhouse with controlled condition was successfully applied, the maternal Longiflorum cultivar'White Fox'performed high susceptable to Fusarium wilt disease while the paternal Asiatic cultivar'Connecticut King'was high resistant, the resistance was successfully inherited to the offspring and segregated in different genotypes. Among the segregated population, majority of the individuals performed different level of resistance to Fusarium wilt disease, the transmission of resistance in the hybrid population reached 70.1%. It can be concluded that although the Fusarium resistance was controlled by quantitative trait loci, some of the genotypes in the F1 hybrids possessed similar resistance when compariing with the paternal material. As a result, first step was achieved in introgression breeding of Fusarium resistance via distant interspecific hybridization.In conclusion, the current research showed that the interspecific hybrids can be successfully obtained via cut-style pollination method and embryo sac culture, cytological observation revealed that some genetypes that showed partial fertility contributed unreduced gametes when backcrossing, and meanwhile, numerical intergenomic recombination, artificial infection of Fusarium oxysporum f.sp. lilii in greenhouse with controlled condition indicated that paternal resistance segregated in the F1 hybbrids, which underlied the recombination of Fusarium resistance genes, were detected in the backcross progeny. Hence, Fusarium resistant breeding of lily through interspecific hybridization became realistic.