基于流式细胞术和K-mer分析的银缕梅属(Parrotia C.A.Mey.)植物基因组大小测定

Estimation of Genome Size of Parrotia C.A.Mey.by Flow Cytometry and K-mer Analysis

金缕梅科(Hamamelidaceae)银缕梅属(Parrotia C.A.Mey.)仅包含银缕梅(Parrotia subaequalis(H.T.Chang)R.M.Hao&H.T.Wei)和波斯铁木(Parrotia persica(DC.)C.A.Mey.)两种落叶阔叶乔木,其中银缕梅是我国华东地区特有的Ⅰ级濒危珍稀保护植物,属东亚第三纪孑遗成分;其姊妹种波斯铁木则间断分布于伊朗北部,属北极第三纪孑遗植物类群。本研究首次利用流式细胞术和K-mer分析方法对银缕梅属两姊妹种的基因组大小进行了测定,建立和优化了以萝卜(Raphanus sativus L.‘Saxa’)为内标、WPB(Woody plant buffer)为细胞核解离液的两种植物单倍体基因组的DNA含量(DNA C值)流式测定的适宜体系,旨在为金缕梅科银缕梅属植物的全基因组测序、基因组学研究、种质资源开发和利用以及物种保育等提供前期基础数据参考;同时也可为金缕梅科其他属、种的基因组大小测定提供借鉴。主要研究结果如下:(1)通过流式测定银缕梅基因组大小约为971.45±13.91 Mb,波斯铁木基因组大小约为890.52±24.69 Mb;(2)K-mer分析估测银缕梅基因组大小为951.70 Mb,杂合率为1.740%,重复序列比例为77.50%;波斯铁木基因组大小为858.50 Mb,杂合率为0.695%,重复序列占74.30%;(3)银缕梅属于高杂合和高重复基因组,波斯铁木则属于微杂合和高重复基因组。本研究的结果为银缕梅属植物后续基于DNA三代高通量测序技术的全基因组测序、组装及去冗余处理等工作提供了重要的数据参考。

The genus Parrotia C.A.Meyer(Hamamelidaceae)contains just two extant deciduous broadleaved trees,Parrotia subaequalis(H.T.Chang)R.M.Hao&H.T.Wei and P.persica(DC.)C.A.Mey.P.subaequalis,classified as‘GradeⅠKey Protected Wild Plant’,is native to eastern China and belongs to East Asian Tertiary relics.Its sibling species,P.persica,is a member of Arctic tertiary relict plant group and disjunct distributed in northern Iran.In this study,we firstly estimated the genome size of two sister species of Parrotia using flow cytometry and K-mer analysis,as well as established and optimized a suitable system for flow cytometric determination of DNA C value of these two plants selecting Raphanus sativus L.‘Saxa’as internal standard and WPB(Woody plant buffer)as nuclear dissociation solution.Our present research aims to provide a fundamental guidance for whole genome sequencing,genomics research,germplasm resource development and utilization,and species conservation of Parrotia in Witch Hazel family and set an important example for the estimation of the genome size of other genera and species of Hamamelidaceae.The main findings are as follows:(1)measurement results of the flow cytometry showed that the estimated genome size of P.subaequalis and P.persica was 971.45±13.91 Mb and 890.52±24.69 Mb,respectively.(2)K-mer analysis estimated that the genome size of P.subaequalis was about 951.70 Mb,the hybridity rate was about 1.740%,and the proportion of repeated sequences was about 77.50%,while for P.persica,the genome size,hybridity rate and the percentage of repeated sequences was 858.50 Mb,0.695%and 74.30%,respectively.(3)The genome of P.subaequalis is classified as a highly heterozygous and highly repeated genome,while P.persica genome is a slightly heterozygous and highly repeated genome.Accordingly,the results of this study provide an important database reference for the follow-up work of whole-genome sequencing,assembly and de-redundancy processing of Parrotia based on DNA third-generation high-throughput sequencing technology.