Citrus sinensis is the most cultivated and economically valuable Citrus species in the world,whose genome has been assembled by three generation sequencings.However,chromosome recognition remains a problem due to the ...Citrus sinensis is the most cultivated and economically valuable Citrus species in the world,whose genome has been assembled by three generation sequencings.However,chromosome recognition remains a problem due to the small size of chromosomes,and difficulty in differentiating between pseudo and real chromosomes because of a highly heterozygous genome.Here,we employ fluorescence in situ hybridization(FISH)with 9 chromosome painting probes,30 oligo pools,and 8 repetitive sequences to visualize 18 chromosomes.Then,we develop an approach to identify each chromosome in one cell through single experiment of oligo-FISH and Chromoycin A3(CMA)staining.By this approach,we construct a high-resolution molecular cytogenetic map containing the physical positions of CMA banding and 38 sequences of FISH including centromere regions,which enables us to visualize significant differences between homologous chromosomes.Based on the map,we locate several highly repetitive sequences on chromosomes and estimate sizes and copy numbers of each site.In particular,we discover the translocation regions of chromosomes 4 and 9 in C.sinensis“Valencia.”The high-resolution molecular cytogenetic map will help improve understanding of sweet orange genome assembly and also provide a fundamental reference for investigating chromosome evolution and chromosome engineering for genetic improvement in Citrus.展开更多
基金supported by the National Natural Science Foundation of China(31970525,31672112)the National Key Research and Development Project of China(2019YFD1001401-GJ03)+4 种基金Advanced Foreign Experts Project(G2021157012L,G20200017071)Cultivating Fund Project of Hubei Hongshan Laboratory(2022hspy002)Fundamental Research Funds for the Central Universities(2662018PY099)from Chinese governmentBreeding of breakthrough new citrus varieties and creation of new germplasms,Science and Technology Planning Project of Sichuan Province(2021YFYZ0023-02)Comprehensive experimental station of middle and late maturing citrus in Chengdu,China Agriculture Research System(CARS-26)。
文摘Citrus sinensis is the most cultivated and economically valuable Citrus species in the world,whose genome has been assembled by three generation sequencings.However,chromosome recognition remains a problem due to the small size of chromosomes,and difficulty in differentiating between pseudo and real chromosomes because of a highly heterozygous genome.Here,we employ fluorescence in situ hybridization(FISH)with 9 chromosome painting probes,30 oligo pools,and 8 repetitive sequences to visualize 18 chromosomes.Then,we develop an approach to identify each chromosome in one cell through single experiment of oligo-FISH and Chromoycin A3(CMA)staining.By this approach,we construct a high-resolution molecular cytogenetic map containing the physical positions of CMA banding and 38 sequences of FISH including centromere regions,which enables us to visualize significant differences between homologous chromosomes.Based on the map,we locate several highly repetitive sequences on chromosomes and estimate sizes and copy numbers of each site.In particular,we discover the translocation regions of chromosomes 4 and 9 in C.sinensis“Valencia.”The high-resolution molecular cytogenetic map will help improve understanding of sweet orange genome assembly and also provide a fundamental reference for investigating chromosome evolution and chromosome engineering for genetic improvement in Citrus.
