昆明植物所揭示限制叶绿体基因组序列进化速率的因素

山茶属是山茶科中包含许多举世闻名经济植物的一个重要类群，包括为人类提供天然保健饮料的茶(Camellia sinensis var. assamica 和C. sinensis var. sinensis)，健康型高级食用植物油的油茶(C. oleifera)以及观赏花卉云南山茶(C. reticulata)、金花茶(C. petelotii)等。然而，迄今为止，山茶属植物遗传学与基因组学研究背景相当薄弱，频繁的种间杂交和多倍化也导致其分类鉴定与系统发育研究十分困难。

叶绿体基因组序列已经被成功地运用于许多物种的分类鉴定和系统关系研究。由于叶绿体基因组较为保守，序列片段变异速率适中，而且叶绿体DNA属于母系遗传，不受遗传重组的影响，因此叶绿体基因组信息的获得不仅对于揭示物种亲缘关系和进行植物系统发育分析有重要意义，还能为植物DNA条形码以及植物叶绿体遗传转化提供大量重要数据信息。

在中国科学院“百人计划”、云南省高端人才项目、云南省海外高层次引进计划和国家自然科学基金的联合支持下，中国科学院昆明植物研究所国家大科学装置中国西南野生生物种质资源库植物种质资源与基因组学研究中心高立志课题组从2006年以来对山茶属中的重要类群进行了系统深入的研究。

该研究按照闵天禄系统，利用Illumina高通量测序平台对山茶属中茶组10个物种或变种、离蕊茶组1个物种、油茶组1个物种和山茶组1个物种的叶绿体基因组进行测序，获得13个叶绿体基因组序列。研究结果发现，山茶属植物叶绿体基因组高度保守，呈现典型的四分体结构，各个物种间IR区的扩张与收缩变异微小。对山茶属系统发育关系最远、充分代表山茶属系统发育关系多样性的4个组的代表种(Camellia sinensis var. assamica, C. oleifera, C. reticulata, C. petelotii)以及分类位置一直存在颇多争议的毛肋茶(C. pubicosta)进行了深入的比较分析，结果表明，无论是重复片段、简单重复序列的类型和数量，还是叶绿体基因组序列间的核苷酸替换、插入、缺失和S/I(substitution/indel)结果，以及基于叶绿体基因组序列重建的山茶属茶组植物系统发育关系，一致支持C. sinensis var. assamica与C. pubicosta具有较近的亲缘关系，认为张宏达系统中将毛肋茶归入茶组是合理的。

通过对叶绿体基因组不同区域的序列分歧度的计算，发现叶绿体基因组序列进化速率不受功能限制而是受区域限制。此外还鉴定、筛选出15个分歧度超过1.5% 的序列作为分子标记，为以后山茶属植物的分类鉴定与系统发育研究提供了重要的信息。

上述研究结果发表于BMC evolutionary Biology上。

原文摘要：

Thirteen Camellia chloroplast genome sequences determined by high-throughput sequencing: genome structure and phylogenetic relationships

Hui Huang, Chao Shi, Yuan Liu, Shu-Yan Mao and Li-Zhi Gao

Background

Camellia is an economically and phylogenetically important genus in the family Theaceae. Owing to numerous hybridization and polyploidization, it is taxonomically and phylogenetically ranked as one of the most challengingly difficult taxa in plants. Sequence comparisons of chloroplast (cp) genomes are of great interest to provide a robust evidence for taxonomic studies, species identification and understanding mechanisms that underlie the evolution of the Camellia species.

Results

The eight complete cp genomes and five draft cp genome sequences of Camellia species were determined using Illumina sequencing technology via a combined strategy of de novo and reference-guided assembly. The Camellia cp genomes exhibited typical circular structure that was rather conserved in genomic structure and the synteny of gene order. Differences of repeat sequences, simple sequence repeats, indels and substitutions were further examined among five complete cp genomes, representing a wide phylogenetic diversity in the genus. A total of fifteen molecular markers were identified with more than 1.5% sequence divergence that may be useful for further phylogenetic analysis and species identification of Camellia. Our results showed that, rather than functional constrains, it is the regional constraints that strongly affect sequence evolution of the cp genomes. In a substantial improvement over prior studies, evolutionary relationships of the section Thea were determined on basis of phylogenomic analyses of cp genome sequences.

Conclusions

Despite a high degree of conservation between the Camellia cp genomes, sequence variation among species could still be detected, representing a wide phylogenetic diversity in the genus. Furthermore, phylogenomic analysis was conducted using 18 complete cp genomes and 5 draft cp genome sequences of Camellia species. Our results support Chang’s taxonomical treatment that C. pubicosta may be classified into sect. Thea, and indicate that taxonomical value of the number of ovaries should be reconsidered when classifying the Camellia species. The availability of these cp genomes provides valuable genetic information for accurately identifying species, clarifying taxonomy and reconstructing the phylogeny of the genus Camellia.

作者：昆明植物所