Lan Hong, Chen Chun-Li, Miao Yin, Yu Chang-Xiu, Guo Wen-Wu, Xu Qiang, Deng Xiu-Xin
Key Laboratory of Horticultural Plant Biology (Ministry of Education), Huazhong Agricultural University, Wuhan, 430070, China.
College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China.
PLoS One. 2016 Mar 15;11(3):e0151512. doi: 10.1371/journal.pone.0151512. eCollection 2016.
Citrus sinensis chromosomes present a morphological differentiation of bands after staining by the fluorochromes CMA and DAPI, but there is still little information on its chromosomal characteristics. In this study, the chromosomes in 'Valencia' C. sinensis were analyzed by fluorescence in situ hybridization (FISH) using telomere DNA and the 45S rDNA gene as probes combining CMA/DAPI staining, which showed that there were two fragile sites in sweet orange chromosomes co-localizing at distended 45S rDNA regions, one proximally locating on B-type chromosome and the other subterminally locating on D-type chromosome. While the chromosomal CMA banding and 45S rDNA FISH mapping in the doubled haploid line of 'Valencia' C. sinensis indicated six 45S rDNA regions, four were identified as fragile sites as doubled comparing its parental line, which confirmed the cytological heterozygosity and chromosomal heteromorphisms in sweet orange. Furthermore, Ag-NOR identified two distended 45S rDNA regions to be active nucleolar organizing regions (NORs) in diploid 'Valencia' C. sinensis. The occurrence of quadrivalent in meiosis of pollen mother cells (PMCs) in 'Valencia' sweet orange further confirmed it was a chromosomal reciprocal translocation line. We speculated this chromosome translocation was probably related to fragile sites. Our data provide insights into the chromosomal characteristics of the fragile sites in 'Valencia' sweet orange and are expected to facilitate the further investigation of the possible functions of fragile sites.
通过荧光染料CMA和DAPI染色后，甜橙染色体呈现出带型的形态分化，但关于其染色体特征的信息仍然很少。在本研究中，以端粒DNA和45S rDNA基因为探针，结合CMA/DAPI染色，通过荧光原位杂交（FISH）对‘伏令夏橙’甜橙的染色体进行分析，结果表明甜橙染色体上有两个脆弱位点，它们共定位于扩展的45S rDNA区域，一个位于B型染色体近端，另一个位于D型染色体亚末端。而‘伏令夏橙’甜橙双单倍体系的染色体CMA带型分析和45S rDNA FISH定位显示有6个45S rDNA区域，其中4个与亲本相比在加倍后被鉴定为脆弱位点，这证实了甜橙的细胞学杂合性和染色体异态性。此外，银染核仁组织区（Ag-NOR）鉴定出在二倍体‘伏令夏橙’甜橙中有两个扩展的45S rDNA区域为活跃的核仁组织区（NORs）。‘伏令夏橙’甜橙花粉母细胞（PMC）减数分裂中四价体的出现进一步证实其为染色体相互易位系。我们推测这种染色体易位可能与脆弱位点有关。我们的数据为‘伏令夏橙’甜橙中脆弱位点的染色体特征提供了见解，有望促进对脆弱位点可能功能的进一步研究。
