| Selection breeding and extension of new cultivars play a key role in citrus industry. Some barriers of citrus breeding, such as nucellar embryony, sterility of some important cultivars, long juvenile phase and high degree of heterozygosity, hindered the selection of breeding parents. As a result cultivar improvement via cross breeding in citrus, one of the best ways for realizing genetic recombination, was severly impeded. Somatic hybridization via protoplast fusion can overcome these obstacles and more than 250 somatic hybrids have been produced in the world so far. Protoplast fusion can be employed to transfer some desirable traits like tolerance or resistance to biotic and abiotic stresses (cold, heat, pest, disease) from donor into target species of high economic value in citrus. In addition, it can combine genetic components from two cultivars with pistil/stamen sterility that can not be obtained via traditional breeding. Somatic hybrids provide valuable germplasms for citrus industry.Triploid cultivars attract the attention from both the producers and the breeders with large and seedless fruits. Triploids are primaril> obtained by hybridization between tetraploids and diploids. Citrus somatic hybrids are allotetraploids having genetic materials from two fusion parents, which differs from the autotetraploids. Hybridization between allotetraploids and diploids will lead to more variations in the progenies that possess three parents' genetic materials. Therefore, the allotetraploids provide more excellent parents for selecting high quality triploid cultivars. Of all the somatic hybrids created to date, about half were derived from fusions between superior scion cultivars, which aims at creating valuable parents for producing seedless and superior triploids through interploidy crossing.The species derived from the natural evolution have wild adaptability to environment, climate and their genetic materials can be transferred to their progenies randomly during man-made crossing or natural crossing. Citrus somatic hybrids incorporate the genetic components from two parents with different genetic backgrounds by artificial measures.However, no reports are available on information concerning the inheritance patterns of different parent's genetic information. The present research is designed to get a hybrid population and investigate the inheritance patterns of fusion parent's genetic information. The populations came from the interploidy cross between somatic hybrid [Hamlin sweet orange (C. sinensis Osbeck) +Rough Lemon (C. jambhiri Luss)] as the pollen parent and the monoembryonic seedy diploid type [No. 4 sexual progeny from Huanongbendizao tangerine (C. reticulata Blanco) (?) X Ichang papeda (C. ichangensis Swingle) ( 8 ) as the female parent. The results are as follows.1. Flow cytometric analysis indicated that 13 out of 92 plants from the sexual crossing were diploids (14.1%) and the rest were triploids (85.9%). The results are the same as that by chromosome counting of root- and shoot-tips.2. The means of leaf length, width and area of sexual progeny are smaller than those of parents and the thickness is similar to that of parents, whereas the menas of stoma length, width and density are intermediate between those of the two parents. Variation ranges of leaf length, width, area, thickness and stoma length, width, density of the sexual progenies are crisscrossing with that of sexual parents. The variation range of other parameters are larger except the leaf thickness. The obvious differences in leaf area and stoma density between diploid and triploid plants result in more variation in the sexual progenies.jf (Chi-Square) test or test of goodness of fit for enumeration data indicated that 1 : 1 segregation was deteced in leaf shape, presence or absence of leaf apex notch and wing leaf shape , in accordance with the distribution rules of the sexual progeny between diploids. If the diploids and triploids are considered as a independent population respectively, by the hypothesis test of the average, notably significant difference was observed in leaf length and stoma length and density between the two populations, together with significant difference in leaf width and stoma width, while the leaf thickness is almost the same. However, there was no obvious gap in leaf length, width, thickens and stoma size, density of single plant between diploids and triploids and their observation value crisscrossed with each other. The stoma of triploid plants were longer while those of diploid plant were more round, the stoma shape index of the former was 1.28(21.71/19.3) and the latter 1.17(21.73/18.15).3. Microscopic observation showed that 97.1% and 96.5% cells underwent somatic mitosis in plants with normal growth vigor and those in weak state respectively, whichdemonstrated that there is possibly no relationship between mitosis with growth vigor by test of independence of enumeration data.4. Totally 14 triploid plants and 1 diploid plant of sexual progenies flowered and 2 triploid plants set fruits. Primary results indicated that the triploids are sterile and the pollen stainability, germinability of the blossomed diploid and triploid plant are much lower than those of their corresponding diploid and tetraploid parents by checking pollen stainability and germinability. Furthermore, 2 out of the 3 fruits in the triploids plants were seedless, and the rest one had only one seed. They had some traits that can be found in their parents.About 26.0% stigmas of floral organ of the flowered sexual progenies degraded, similar to that of one of the fusion parents (Rough Lemon). Of the progenies, the ratios of normal flowers in the triploids and the diploids are nearly the same, with 73.0% in triploid progenies with little variation range and 76.0% in diploids, which is intermediate between their parents. The flowers of the sexual progeny were pink at earlier stage, similar to Ichang papeda and Rough Lemon. The pink flowers became white as the flowers grew, the same as Huanong bendizao tangerine and Hamlin sweet orange, like parent of IB4 and HR. respectively. When it bloomed, the anthers of triploid were white and shrunk, the same as C. unshiu Marc, while those of the diploid were also shrunk and light yellow in color.5. Thirty-one 10-mer primers have been selected from the 154 ones, which could give rise to clear bands, have good polymorphisms and are reproducible. They were used to identify the hybrids and analyze the inheritance patterns. All of the 92 sexual progenies were confirmed as hybrids with the genetic message of three parents regardless of diploid or triploid by RAPD and SSR .The total DNA of the progenies and their parents was amplified using the 31 polymorphic primer by the means of RAPD. -ï¿¡ analysis of the amplification results showed that markers specific to either parent were transmitted to the progenies randomly. If the diploid and the triploid plants are considered as independent populations respectively, the average of common markers presented in either of the fusion parents namely Hamlin sweet orange or Rough Lemon and the female parent in the diploid population is larger than that in the triploid population, while the number of markers common and unique to three parents IB4, Hm and RL, markers shared by the fusion parents, the total number of parent markers present in individual plant and polymorphism degree are larger in the triploid population than in the diploid one. However, hypothesis test of average revealed that there is no significant difference in theaverage markers from parents between the diploids and the tripoids, which further supported the random distribution of parents' specific markers in progenies and absence of relationship between the ploidy and number of parent's markers. In additional, the variation range was reduced concerning parents' total markers, specific makers, common markers and polymorphism degrees among the diploid plants and the triploid plants, which means improved the uniformity.The genetic distance of the 92 progeny plants and the parent is calculated by software NTSYS pc(version 2.10e) and finally a dendrogram was constructed by UPGMA. The sexual progenies and the parents can be clustered into 3 groups. The first group is clustered with one of the fusion parents (Hamlin sweet orange), including 15 sexual progeny plants (16%) The second group is clustered with female plant IB4, including 16 sexual progeny plants, of which 7 plants are diploid, and the third group is clustered with the somatic hybrid as male parent (Hamlin sweet + Rough Lemon), including 61 sexual progeny plants and Rough Lemon (66%). The plants of the third group can be sub-divided into 3 groups and 15 sexual progeny plants were clustered with Rough Lemon. In general, crossing with diploid type using allotetraploid citrus somatic hybrid as the pollen parent, most sexual progenies had closer genetic relation with the paternal.6. RFLP analysis of some progeny plants and the related parents with polymorphic restriction endonucleases /cytoplasmic probe combinations demonstrated that regardless of mitochondrial probe or chloroplast probe, the hybridization profiles of the sexual progenies were identical to those of female parent, indicating that the cytoplasmic genomes of citrus followed the maternal inheritance, the same as that of other plant species.
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