caa a22 4500 445840277 CHVBK 20180317145341.0 cr unu---uuuuu 170323e20110701xx s 000 0 eng 10.1007/s00709-010-0200-2 doi (NATIONALLICENCE)springer-10.1007/s00709-010-0200-2 Effect of ploidy increase on transgene expression: example from Citrus diploid cybrid and allotetraploid somatic hybrid expressing the EGFP gene [Elektronische Daten] [Shi-Xiao Xu, Xiao-Dong Cai, Bin Tan, Ding-Li Li, Wen-Wu Guo] Polyploidization is an important speciation mechanism for all eukaryotes, and it has profound impacts on biodiversity dynamics and ecosystem functioning. Green fluorescent protein (GFP) has been used as an effective marker to visually screen somatic hybrids at an early stage in protoplast fusion. We have previously reported that the intensity of GFP fluorescence of regenerated embryoids was also an early indicator of ploidy level. However, little is known concerning the effects of ploidy increase on the GFP expression in citrus somatic hybrids at the plant level. Herein, allotetraploid and diploid cybrid plants with enhanced GFP (EGFP) expression were regenerated from the fusion of embryogenic callus protoplasts from ‘Murcott' tangor (Citrus reticulata Blanco × Citrus sinensis (L.) Osbeck) and mesophyll protoplasts from transgenic ‘Valencia' orange (C. sinensis (L.) Osbeck) expressing the EGFP gene, via electrofusion. Subsequent simple sequence repeat (SSR), chloroplast simple sequence repeat and cleaved amplified polymorphic sequence analysis revealed that the two regenerated tetraploid plants were true allotetraploid somatic hybrids possessing nuclear genomic DNA of both parents and cytoplasmic DNA from the callus parent, while the five regenerated diploid plants were cybrids containing nuclear DNA of the leaf parent and with complex segregation of cytoplasmic DNA. Furthermore, EGFP expression was compared in cells and protoplasts from mature leaves of these diploid cybrids and allotetraploid somatic hybrids. Results showed that the intensity of GFP fluorescence per cell or protoplast in diploid was generally brighter than in allotetraploid. Moreover, same hybridization signal was detected on allotetraploid and diploid plants by Southern blot analysis. By real-time RT-PCR and Western blot analysis, GFP expression level of the diploid cybrid was revealed significantly higher than that of the allotetraploid somatic hybrid. These results suggest that ploidy level conversion can affect transgene expression and citrus diploid cybrid and allotetraploid somatic hybrid represents another example of gene regulation coupled to ploidy. Springer-Verlag, 2010 Citrus nationallicence Confocal laser scanning microscopy nationallicence Green fluorescent protein nationallicence Ploidy level nationallicence Somatic hybridization nationallicence CAPS : Cleaved amplified polymorphic sequence nationallicence CLSM : Confocal laser scanning microscope nationallicence cpDNA : Chloroplast DNA nationallicence EGFP : Enhanced GFP nationallicence GFP : Green fluorescent protein nationallicence MT medium : Murashige and Tucker (1969) nationallicence mtDNA : Mitochondrial DNA nationallicence SSR : Simple sequence repeat nationallicence Xu Shi-Xiao Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut Cai Xiao-Dong Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut Tan Bin Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut Li Ding-Li Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut Guo Wen-Wu Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut Protoplasma Springer Vienna 248/3(2011-07-01), 531-540 0033-183X 248:3<531 2011 248 709 https://doi.org/10.1007/s00709-010-0200-2 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00709-010-0200-2 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Xu Shi-Xiao Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut NATIONALLICENCE 700 1- Cai Xiao-Dong Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut NATIONALLICENCE 700 1- Tan Bin Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut NATIONALLICENCE 700 1- Li Ding-Li Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut NATIONALLICENCE 700 1- Guo Wen-Wu Key Laboratory of Horticultural Plant Biology (Ministry of Education); National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China aut NATIONALLICENCE 773 0- Protoplasma Springer Vienna 248/3(2011-07-01), 531-540 0033-183X 248:3<531 2011 248 709 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer