Separation Of Citrus Fruit Sector Chimeras And Genetic Analysis Of Two Graft Chimeras

Selection of bud mutation is one of the most important methods for Citrus breeding. Generally, citrus fruit sector chimeras and graft chimeras were considered to belong to bud mutation and could be used as genetic resources for cultivar improvement. In the present research, a 5-year investigation was carried out to study the frequency of citrus fruit sector chimeras. And some chimeras were separated and purified by in vitro technique as the candidates for selecting new cultivars. Two graft chimeras were genetically evaluated, their layer constructions were studied. The main results are as follows:1. During the years of 2002-2006, frequency of citrus fruit sector chimeras were investigated in 6 citrus cultivars in an orchard of Zigui county, Hubei province. The cultivars included'Newhall' navel orange(Citrus sinensis Osbeck),'Changhong' navel orange(C. sinensis Osbeck),'Cara Cara' navel orange(C. sinensis Osbeck), navel blood orange(C. sinensis Osbeck),'Carter' orange(C. sinensis Osbeck) and'Ponkan'(C. reticulata Blanco). During the years of 2003-2006, same work was conducted in 3 citrus cultivars in the orchards of Xunwu county, Jiangxi province. The cultivars included'Newhall' navel orange,'Navelina' navel orange(C. sinensis Osbeck) and'Bonanza' navel orange(C. sinensis Osbeck). During the years of 2004-2006, same investigation of 2 citrus cultivars in the orchards of Xingshan county, Hubei province was carried out. The cultivars included'Newhall' navel orange and'Jincheng' sweet orange(C. sinensis Osbeck). The results indicated that the frequency of the fruit sector chimeras of nine citrus cultivars' varied from 0.02% to 3.97%. The frequency in'Cara Cara' navel orange(3.97%) is the highest and that in'Ponkan' is the lowest(0.02%). Frequency varied with genotypes, environment and cultivation methods. Different types of sector mutations were observed in chimeric citrus fruits, out of them alterations of sector rind color were predominatedly observed, as shown by brown, green, yellow, dark red, dark orange, pink sectors. Another common mutation was the thicker or thinner rind sector than the normal.2. A total of 180 chimeric fruits were collected from 9 cultivars described above. 30 seeds and 5000 aborted ovules from the fruit chimeras were cultured and 2078 plantlets were recovered from them. Among them, 1508 plantlets were recovered from the seeds and aborted ovules beneath the chimeric part of each fruit and 570 plantlets were from the normal sectors. Among 600 plantlets which were recovered from 50 fruit chimeras with thicker rind, 3 tetraploid plants of'Newhall' navel orange, 4 of'Navelina' navel orange and 2 mixploid plants from'Newhall' navel orange were obtained after flow cytometry confirmation. In total, 1000 plantlets were transferred to the greenhouse and field and which will be the candidates for new cultivars selections.3. Two new strains of citrus from graft chimeras were collected as were named'Zaohong' navel orange(Citrus sinensis Osbeck+Citrus unshiu Marc.) and'Hongrou Taoye' sweet orange(Citrus sinensis Osbeck+Citrus unshiu Marc.) respectively.'Zaohong' navel orange arose at the junction area where a'Robertson' navel orange(C. sinensis Osbeck) scion was top-worked onto Satsuma mandarin(C. unshiu Marc).'Hongrou Taoye' sweet orange was from the junction where a'Taoye' sweet orange(C. sinensis Osbeck) scion was top-worked onto Satsuma mandarin. These two graft chimeras and their donors were evaluated by morphological, cytological, biochemical and molecular markers. Characters of the'Zaohong' navel orange and'Hongrou Taoye' sweet orange, such as juice sacs of fruits, stoma length, which were determined by L1 cell layer, were similar to Satsuma mandarin. While other traits, including leaf index, fruit shape, navel, pollen morphology and fertility, the number of seeds, the color and aroma of the rind determined by L2 cell layer, were the same as'Robertson' navel orange and'Taoye' sweet orange respectively. The carotenoid extract of the flesh of'Zaohong' navel orange,'Hongrou Taoye' sweet orange and their donor plants was studied by using high performance liquid chromatography(HPLC). The results indicated that'Zaohong' navel orange and'Hongrou Taoye' sweet orange had the same carotenoids profile of Satsuma mandarin, in whichβ-cryptoxanthin accumulated predominantly in the juice sacs in mature fruit. SSR and cpSSR analysis showed that both nuclear and chloroplast genomes of'Zaohong' navel orange and'Hongrou Taoye' sweet orange were composed from both of their donor plants respectively. Based on these facts,'Zaohong' navel orange was a periclinal chimera consisting of L1 derived from Satsuma mandarin and L2/L3 from'Robertson' navel orange.'Hongrou Taoye' sweet orange was a periclinal chimera consisting of L1 derived from Satsuma mandarin and L2/L3 from'Taoye' sweet orange. They combined the elite traits of their donor plants and genetically stable, they could be commercially acceptable chimeric cultivars for the fresh market of citrus.4. SSR primer TAA27 was used to verify the seedlings derived from the nucellar embryo of'Zaohong' navel orange and'Hongrou Taoye' sweet orange. The results showed that the band model of the plants were the same as the L2 donors. And it is concluded that the nucellar embryo in citrus is derived from the cell in L2 layer.