甜橙与酸橙体细胞杂种核质组成鉴定(英文)

采用流式细胞术(flow cytometry, FCM)、简单重复序列(simple sequence repeat, SSR)和酶切扩增多型性序列(cleaved amplified polymorphic sequence, CAPS)等技术分析酸橙(Citrus aurantium L. )叶肉原生质体和甜橙(C. sinenis Osbeck cv. Shamouti)胚性愈伤组织原生质体电融合再生的体细胞杂种。FCM研究结果表明,所有的体细胞杂种植株荧光强度是二倍体对照的2倍,说明所分析的植株为四倍体。用SSR和CAPS分析了体细胞杂种的核质遗传组成,在试验的4对SSR引物中,有2对能区分开融合亲本。在2对引物中,体细胞杂种植株包含双亲的全部特异带,表明它们为异核杂种。通用引物扩增结合限制性内切酶酶切能鉴别融合亲本,在具有多型性的引物/酶组合中,所有体细胞杂种的线粒体和叶绿体DNA带型与胚性亲本(甜橙)完全一样。结果表明体细胞杂种核基因组来自双亲,而胞质基因组来自悬浮系亲本。讨论了所用技术的特点、柑橘四倍体体细胞杂种核质遗传规律及本组合体细胞杂种的应用。

Characterization of Nuclear and Cytoplasmic Compositions of Somatic Hybrid Plants Between Sweet Orange and Sour Orange

Abstract In the present research, flow cytometry (FCM), simple sequence repeat (SSR) and cleavedamplified polymorphic sequence (CAPS) were employed to analyze somatic hybrid plants derived from electrofusion between embryogenic protoplasts of sweet orange (Citrus sinensis Osbeck cv. Shamouti) and leaf-derived protoplasts of sour orange (C. aurantium L). FCM showed that all of the somatic hybrid plants had two-fold fluorescence intensity values of the diploid control, indicating that they were tetraploids. SSR and CAPS were used to characterize the compositions of nuclear and cytoplasmic genomes of the somatic hybnd plants. As for SSR four primer pairs were tried and two showed polymorphisms between the fusion parents. With both pnmer pairs the somatic hybrid plants encompassed DNA bands from both parents, indicating that they were heterokaryonic hybrids. Amplification with some universal primers, followed by digestion with restriction endonucleases, could distinguish the fusion parents from each other. As far as mitochondrial and chloroplast DNA compositions were concerned the somatic hybrid plants shared the same banding patterns as the embryogenic parents for all of the polymorphic primer/enzyme combinations. The results herein demonstrated that the somatic hybrid plants inherited their nuclear genome from both fusion parents, whereas the cytoplasmic genomes were possibly only contributed by the embryogenic parent. Merits of the analytical methods and nuclear and cytoplasmic inheritance of citrus tetraploid somatic hybrids, together with their application, are discussed herein.