关键词: 柑橘, 叶绿体基因组InDel, 叶绿体基因组SSR, 遗传进化, 种质资源

Abstract:

【Objective】The genetic diversity analysis of the chloroplast genome was performed to reveal the genetic evolution of citrus from the cytoplasmic level, so as to provide a reference for the collection, evaluation and utilization of citrus germplasm resources. 【Method】Taking the hypervariable region of Citrus sinensis chloroplast genome as reference, De Novo assemblies were carried out with next-generation sequencing data of representative Citrus and its related genera materials. The assembled sequences were aligned to discover site-variations, and cpInDel markers were designed on the basis of variation sites. SSR sites composed of two or more nucleotide repeats were located in the chloroplast genome of sweet orange, and SSR sites with length variations were adopted to design cpSSR markers. The newly developed markers and reported cpSSR markers were used to detect genetic diversity in 48 samples of Citrus and its relatives that are representative in taxonomy and/or origin through band typing and clustering analysis. 【Result】4 cpInDel markers and 13 cpSSR markers were developed successfully and they could produce clear polymorphic bands in all tested samples. The band pattern of cpIndel was relatively simple. Single marker could distinguish a specific genus or several genera. At species and/or above taxa level, the cpInDel markers gave more convenient and accurate discrimination effect. Similar clustering results were obtained with cpInDel and cpSSR markers, and minor differences were found in the relationship between the lower-rank taxa and the classification of a few genotypes. The cytoplasmic origins of P. trifoliate hybrids were found to be different from P. trifoliata, and those of lemon, limonia and lime hybrid were different from C. medica by both markers. Wangcang zhoupigan, a representative variety of C. speciosa, had a different cytoplasmic origin from other mandarin varieties. Ziyang xiangcheng and Korean xiangcheng, both classified as C. junos, might have different cytoplasmic origin. cpSSR markers classify mandarin, sweet orange and sour orange, ichang papeda and honghe papeda into three clusters, representing C. reticulata, C. sinensis and Papeda, while cpInDel markers showed that they were closely related and classified into one cluster. Indian wild mandarin were clustered into C. medica by cpInDel, and were put into a single cluster by cpSSR. 【Conclusion】Molecular marker analysis of chloroplast genome could reveal the genetic relationship of Citrus and its related genera and discover a different relationship from the view of nuclear genome. However, if combined with markers from nuclear genome, more comprehensive and accurate indentification and phylogenetic relationship revealing should be obtained for the citrus germplasm resources.

Key words: Citrus, chloroplast genome InDel, chloroplast genome SSR, genetic evolution, germplasm