Somatic Hybridization Of Hong Kong Kumquat With Early Flowering Lemon And Preconcious Trifoliate Orange And Molecular Identification Of Regenerated Plants

Citrus occupies important position in horticultural economics in our country with the high yield and good quality being its core competitiveness.Longer juvenility is one of the main barriers for citrus breeding,and the production of short juvenility new germplasm has always been an important goal in citrus breeding.Cell fusion can achieve distant hybridization,and by using protoplasts isolated from short juvenile resources for protoplasts fusion with protoplasts of other diploid species,is expected to shorten the breeding period.Based on this idea,this study aims to obtain new short juvenile germplasm with the advantages of comprehensive parents,and provides new species for citrus breeding and basic research by fusion of short juvenile resources such as Hong kong kumquat with Early flowering lemon and preconcious trifoliate orange.1.Hong kong kumquat(callus parent)+ Early flowering lemon(leaf parent): 16 regenerated plants were survived finally,from 13 different fusion events.Flow cytometry showed that 12 of the regenerated plants(from different fusion events)were diploid and one strain was hexaploid.The SSR and cpSSR molecular markers were used to identify the cytoplasm and nuclear genome(14 pairs of different primers).The results showed that the nuclear genome and chloroplast genome of 12 diploid regenerated plants were obtained from the Early flowering lemon and the mitochondrial genome is derived from the callus of the Hong kong kumquat,that is,the 12 diploid plants are cytoplasmic hybrids while the hexaploid plants need to be further identified.2.Hong Kong Kumquat(callus parent)+ Preconcious trifoliate orange(leaf parent): The enzymatic digestion of the modified enzyme was used to obtain the protoplasts from preconcious trifoliate orange,Still using the laboratory commonly used methods for fusion and regeneration,the combination ultimately only get regenerative callus by multiple fusion experiments.The ploidy level of the regenerative callus was tetraploid by flow cytometry analysis and the plants were not successfully regenerated,possibly due to the relationship between Hong kong kumquat and Precocious trifoliate orange is too far,fusion cells are not easy to regenerate plants.In conclusion,a number of diploid cytoplasm and hexaploid plants were obtained by somatic hybridization.These plants may serve as useful materials for citrus cell engineering basic research.