Research On Construction Of Fingerprint And Reproductive Biology Of Zantedeschia

Genus Zantedeschia is a member of family Araceae. There are7species and2subspecies inthis genus. According to growth habit, they are divided into two sections. Section1is calledEvergreen, another section is named Deciduous. Evergreen section has only one popular species,Z. aethiopica,with the advantages of good cold-tolerance and disease-resistant. Cultivars inDeciduous section are called Calla Lily because they are rich in color and types. Calla Lilycultivars are the current international popular main products. There are more than60cultivarswith almost worldwide use now among more than150registered commercial cultivars. Mostcultivars in this genus were bred by the companies and research institutes from United States,New Zealand and Netherlands.In this research88cultivars resources were used as materials. Based on morphologyobservation and technology methods such as, ISSR-PCR molecular mark, paraffin slice, andpollen micro-observation and cultivars breeding, the morphology DNA fingerprint andreproductive biology were studied. The important morphological characters were observed anddescribed in detail, the guidelines for the Distinctess, Uniformity and Stability （DUS） test ofGenus Zantedeschia were filled out based on the analysis of distinctive, uniform and stabilitycharacteristics. Based on ISSR-PCR molecular mark results, classing standard for4cultivarsgroup were advanced. Megaspore and microspore and the development of gametophytes wereanalyzed. Microscopic morphological characteristics of pollen were observed, the pollen viabilityand germination characteristics were verified. The main conclusions are as follows:1. The leave blade type can be divided into lanceolate, ovate, triangular and hastate. Spathecolors have7types such as white, yellow, orange, pink, red, purple, and bicolor. Spathe viewedfrom top divided into three types: acute, obtuse and rounded.2. All samples have a good consistency, stability and specificity. After the consistencyanalysis of morphological traits,39characters were carried out for DUS test and with goodconsistency compared with UPOV test guidelines. Guidelines for the conduct of tests forDistinctness, Uniformity and Stability Genus Zantedeschia were formulated. The standarddraft for approval was send up to Ministry of Agriculture.3. Orthogonal experimental design was conducted to establish the reaction system.The total25μl reaction solution includes2.5μl10×PCR buffer，2.0mmol/l Mg²⁺,0.20mmol/l dNTPs，1.5UTaq enzyme,0.25μmol/l primer and1.0ng/μl DNA template.15primers were selected from96pairs for ISSR-PCR reaction. These primers amplified690loci.435loci among them werepolymorphic. Average per pair of primers amplified29polymorphic loci out of46loci. Thepolymorphism ratio was63.04%. 4. Using the UPGMA cluster based on molecular markers of ISSR-PCR results,88samplescould be divided into two groups. Zantedeschia aethiopica and Calla lily at similar coefficient of0.15. At similar coefficient of0.29,87Calla cultivars could be divided into3sub-grops, thepot-flower sub-group, cut-flower sub-group and pot-cut flower sub-group. In each sub-groupthere are white, yellow, orange, red and purple color types. This result was similar withmorphological classification.5. Six core primer were selected to establish the fingerprint of Zantedeschia cultivars. Thefingerprint model of samples was drawn.6. The megasporogensis, microsporogensis and the development of female and malegametophytes of Zantedeschia ‘Majestic Red’ were observed through traditional paraffin sections.It’s ovule is anatropous, unitegmic, and crassinucellar with integument tapeta. The megasporemother cell meiosis to form a linear megaspore tetrad. Among them, The chalazal end megasporedevelops into the embryo sac mother cell，which developed to Polygonum type embryo-sac，another3megaspores degenerated. Each flower had2-3stamens, with butterfly-shaped pollensacs，ach side with2small sporangial. The wall of anther was completely developed, whichconsisted of four layers: epidermis, endothecium, middle layers and tapetum. The tapetum isdeformation type. The cytokinesis of microspore mother cell during meiosis was ofsimultaneaous type. Microspore tetrads are mainly cross. Pollen grains had2-celled when shed.During the Microspore Tetrad， tapetum cells Degradated in advance，so the released smallspores couldn’t develop normally7. According to scanning electron microscope observation, pollens among all samples aresimilar in size and shape, prolate shape. The length of polar axis and equatorial axis was for28.8～39.5μ m and21.9～32.3μ m.the P/E values range from1.14～1.43. Pollen extine wassmooth without aperture, pollen sculpture was not obvious.8. At25℃in the dark condition, pollen germination ratio and pollen tube length grew to thehighest value within24hours, especially in early9h. The suitable medium for cultivation ofpollen viability testing was mixed with another10%sucrose and0.01%H3BO3.