High quality genomes produced from single MinION flow cells clarify polyploid and demographic histories of critically endangered Fraxinus (ash) species

With populations of threatened and endangered species declining worldwide, efforts are being made to generate high-quality genomic records of these species before they are lost forever. Here, we demonstrate that data from single Oxford Nanopore Technologies (ONT) MinION flow cells can, even in the absence of highly accurate short DNA-read polishing, produce high-quality de novo plant genome assemblies adequate for downstream analyses, such as synteny and ploidy evaluations, paleodemographic analyses, and phylogenomics. This study focuses on three North American ash tree species in the genus Fraxinus (Oleaceae) that were recently added to the International Union for Conservation of Nature (IUCN) Red List as critically endangered. Our results support a whole genome triplication at the base of the Oleaceae as well as a subsequent whole genome duplication shared by Syringa, Osmanthus, Olea, and Fraxinus. Finally, we demonstrate the use of ONT long-read sequencing data to reveal patterns in ..., Fraxinus pennsylvanica vouchers and material for DNA extraction were collected from Point Gratiot Park in Dunkirk, New York, USA. F. americana and F. nigra vouchers and material for DNA extraction were collected from the College Lodge Forest in Brocton, New York, USA. Samples were sequenced on an Oxford Nanopore Technologies’ GridION instrument utilizing MinION flowcells (version R9.4). Genome assemblies were generated using Flye and reduced to haploid assemblies using Purge Haplotigs. Genomes were additionally scaffolded using RagTag and the recent Fraxinus pennsylvanica reference assembly as a scaffolding reference. All annotations were generated with GeMoMa. ,
Guppy v5.0.11
Nanoplot v1.38.0
Nanostat v1.5.0
Flye v2.8.3
Flye v2.9
QUAST v5.0.2
BUSCO v5.4.4
GeMoMa v1.9
AGAT v1.0.0
Evidential Gene
RepeatModeler v2.0.1
RepeatMasker v4.0.1
RagTag v2.1.0
MCScan (https://github.com/tanghaibao/jcvi/wiki/MCscan-(Python-version))
HapPy (https://github.com/AntoineHo/HapPy)

SAMtools v1.14
Purge Haplotigs v1.1.1
Minimap2 v2.20
bcftools version 1.14
PSMCR (https://github.com/emmanuelparadis/psmcr)

Orthofinder v2.5.4
Ksrates v1.1.3
, # High quality genomes produced from single MinION flow cells clarify polyploid and demographic histories of critically endangered *Fraxinus* (ash) species

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The following data are genome assemblies and annotations generated in this study. In the case of v0.2.1, 1.4.1, and 1.1, the genomes were previously published and we generated new annotations using GeMoMa.

## Description of the data and file structure

Figure_1_Data.zip #contains data for Figure 1
\|-- Figure_1a #MCScan files for generating the karyotype plot in Figure 1a
\| |-- *.bck #file generated by MCScan for pairwise syneny search
\| |-- *.bed #CDS sequence coordinates on assembly
\| |-- *.cds #CDS sequence fasta file
\| |-- *.des #file generated by MCScan for pairwise syneny search
\| |-- *.prj #file generated by MCScan for pairwise syneny search
\| |-- *.sds #file generated by MCScan for pairwise syneny search
\| |-- *.ssp #file generated by MCScan for pairwise syneny search
\| |-- *.suf #file generated by MCScan for p...

全球受威胁与濒危物种种群正持续衰退，科研人员正全力赶在这些物种永久消失前，获取其高质量基因组记录。本研究证实，仅依靠牛津纳米孔技术（Oxford Nanopore Technologies, ONT）MinION流动池产生的数据，即便不进行高精度短DNA读段校正，也可产出高质量的植物从头基因组组装结果，足以支撑后续的同线性分析、倍性评估、古种群历史分析及系统基因组学研究。本研究聚焦于3个北美梣属（*Fraxinus*，木犀科Oleaceae）树种，这些物种近期被国际自然保护联盟（International Union for Conservation of Nature, IUCN）红色名录列为极危物种。研究结果支持木犀科基部发生过一次全基因组三倍化事件，且丁香属（*Syringa*）、木犀属（*Osmanthus*）、木犀榄属（*Olea*）及梣属（*Fraxinus*）共同经历了一次后续的全基因组复制事件。最后，本研究展示了利用ONT长读段测序数据揭示...的模式。灰毛梣（*Fraxinus pennsylvanica*）的凭证标本及DNA提取材料采自美国纽约州敦刻尔克的格拉蒂奥特角公园（Point Gratiot Park）；美国白梣（*F. americana*）与黑梣（*F. nigra*）的凭证标本及DNA提取材料采自美国纽约州布罗克顿的学院洛奇森林（College Lodge Forest）。样本采用牛津纳米孔技术的GridION测序仪，搭配MinION流动池（版本R9.4）完成测序。基因组组装通过Flye完成，并借助Purge Haplotigs将组装结果降为单倍体组装；后续利用RagTag及最新发布的灰毛梣参考基因组作为锚定参考，对基因组进行支架挂载。所有基因组注释均通过GeMoMa生成。
所用分析软件及版本如下：Guppy v5.0.11、Nanoplot v1.38.0、Nanostat v1.5.0、Flye v2.8.3、Flye v2.9、QUAST v5.0.2、BUSCO v5.4.4、GeMoMa v1.9、AGAT v1.0.0、Evidential Gene、RepeatModeler v2.0.1、RepeatMasker v4.0.1、RagTag v2.1.0、MCScan（https://github.com/tanghaibao/jcvi/wiki/MCscan-(Python-version)）、HapPy（https://github.com/AntoineHo/HapPy）、SAMtools v1.14、Purge Haplotigs v1.1.1、Minimap2 v2.20、bcftools v1.14、PSMCR（https://github.com/emmanuelparadis/psmcr）、Orthofinder v2.5.4、Ksrates v1.1.3。
# 基于单MinION流动池产出的高质量基因组解析极危梣属（*Fraxinus*，梣树）物种的多倍化与种群历史

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以下为本研究产出的基因组组装结果及注释文件。针对版本为v0.2.1、1.4.1及1.1的基因组，其此前已发表，本研究通过GeMoMa重新生成了对应的注释信息。

## 数据与文件结构说明
Figure_1_Data.zip # 包含图1的相关数据
|-- Figure_1a # 用于生成图1a核型图谱的MCScan分析文件
| |-- *.bck # MCScan用于成对同线性搜索生成的文件
| |-- *.bed # 组装序列上的CDS序列坐标文件
| |-- *.cds # CDS序列FASTA文件
| |-- *.des # MCScan用于成对同线性搜索生成的文件
| |-- *.prj # MCScan用于成对同线性搜索生成的文件
| |-- *.sds # MCScan用于成对同线性搜索生成的文件
| |-- *.ssp # MCScan用于成对同线性搜索生成的文件
| |-- *.suf # MCScan用于生成p...的文件