濒危植物短叶黄杉叶绿体全基因组特征及松科植物系统发育分析

Endangered Species of Pseudotsuga brevifolia Chloroplast Genome Characteristics and Pinaceae Species Phylogenetic Analysis

为确定短叶黄杉(Pseudotsuga brevifolia)叶绿体基因组结构大小、特征以及基因组成情况,明确其与同科物种叶绿体基因组差异性,并揭示松科植物潜在的变异序列,探究系统发育规律。本研究以二代测序为基础,对测序结果进行组装、注释,得到短叶黄杉叶绿体全基因组,分析了其基本结构、基因组成、SSR位点等数据,并与其他松科植物叶绿体基因组进行比对分析、IR扩张收缩分析与系统发育分析。结果表明:短叶黄杉叶绿体基因组全长122348 bp,呈典型的四分体结构,LSC区长度为66627 bp,SSC区长54863 bp,IR区长度仅为858 bp,一共注释到114个基因,其中有73个蛋白编码基因,37个tRNA基因和4个rRNA基因,5个基因具有2~3个拷贝且都为tRNA基因,且含有内含子的基因共有7个。SSR位点中,单核苷酸重复共有15个,占所有SSR的52%,2~5核苷酸重复分别为6、4、3和1个,主要为AT类型重复。叶绿体基因组序列比对发现,短叶黄杉与黄杉的相似度最高,多数物种在IR区、tRNA和rRNA基因区域的一致度相对较高,最为保守;包括短叶黄杉在内的其他9个物种中,非编码区的遗传变异比编码区更为普遍。IR边界扩张与收缩分析结果表明:9个物种LSC/IRb边界处均为rpl23基因,IR区均发生了明显的收缩,因此基因分布较少且单一。云杉属的两种植物IRb区没有基因分布,云南松IRb区分布有两个基因,其IR区较其他物种发生了扩张。短叶黄杉IR区内分布的基因为trnM,其余物种IR区内的基因均为trnI,且云杉属两个物种的IR/SC边界与松科其他物种差异较大。构建叶绿体全基因组NJ系统发育树显示,松科10个属16个物种主要被分为3大类,欧洲落叶松(Larix decidua)与黄杉属亲缘关系最近。本研究首次报道了短叶黄杉的叶绿体基因组结构与特征,阐明了松科不同属物种间叶绿体基因组的结构差异,明确了短叶黄杉的分类位置,为濒危植物短叶黄杉的遗传育种、分类保护以及松科植物的物种鉴定、种间进化研究和系统发育重建提供了数据基础与参考依据。

To determine the size,characteristics and genetic composition of the chloroplast genome of P.brevifolia,clarify the differences between the chloroplast genome and that of other species in the same family,and to reveal the potential variation sequences of Pinaceae species,and explore the phylogenetic rules.Based on the next-generation sequencing get the chloroplast complete genome of P.brevifolia,we analyzed its basic structure,SSR site,compared chloroplast genome with other Pinaceae species,IR expansion and contraction analysis and phylogenetic analysis to reveal the potential variation sequence of Pinaceae species,explore the phylogenetic law.The results showed that the total length of the chloroplast genome of P.brevifolia was 122348 bp,LSC area length was 66627 bp,SSC area length was 863 bp,IR length was 858 bp.A total of 114 encoded genes which were in the chloroplast genome of P.brevifolia,including 73 protein-coding genes,37 tRNA genes,and 4 rRNA genes.5 genes with 2 to 3 copies(all belonged to tRNA genes),otherwise,7 genes has intron.There were 15 single nucleotide repeats in SSR sites,accounting for 52%of all SSR,and 2~5 nucleotide repeats were 6,4,3 and 1,respectively,and were mainly AT type repeats.The chloroplast genome sequence alignment showed that the similarity between the P.brevifolia and P.sinensis was the highest,and most consistency of most species in the IR,tRNA and rRNA gene regions was relatively high and the most conserved.Genetic variation in noncoding regions was more common in P.brevifolia and other species.The results of IR/SC boundary expansion and contraction analysis showed that the LSC/IRb boundaries of nine species were all rpl23 genes,and the IR regions were contracted,so the gene distribution was less and single.There was no gene distribution in the IRb area of the two plants of Picea,and there were two genes in the IRb area of Pinus yunnanensis,and the IR area of P.yunnanensis was expanded compared with other species.The genes distributed in the IR region of P.brevifolia were trnM,and the genes in the IR region of the other species were all trnl,and IR/SC boundaries of the two species of Picea were significantly different from those of other species of the Pinaceae family.Construction of chloroplast genome NJ phylogenetic tree showed that 16 species of Pinaceae family were divided into 3 categories.Larix decidua was most closely related to the Pseudotsuga genus.This study first reported the chloroplast genome structure and characteristics of P.brevis,clarified the structural differences of the chloroplast genome among the different genera of P.brevifolia,and defined the taxonomic location of P.brevifolia which provided data basis and reference for genetic breeding,taxonomic protection,species identification,interspecific evolution and phylogenetic reconstruction of endangered plants in Pinaceae.