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快速有效的方法来开发孤儿物种的基因组资源:在薰衣草中的应用。

Quick and efficient approach to develop genomic resources in orphan species: Application in Lavandula angustifolia.

机构信息

ITEIPMAI, Chemillé-en-Anjou, France.

US1279 Etude du Polymorphisme des Génomes Végétaux (EPGV), INRA, Université Paris-Saclay, Evry, France.

出版信息

PLoS One. 2020 Dec 11;15(12):e0243853. doi: 10.1371/journal.pone.0243853. eCollection 2020.

Abstract

Next-Generation Sequencing (NGS) technologies, by reducing the cost and increasing the throughput of sequencing, have opened doors to generate genomic data in a range of previously poorly studied species. In this study, we propose a method for the rapid development of a large-scale molecular resources for orphan species. We studied as an example the true lavender (Lavandula angustifolia Mill.), a perennial sub-shrub plant native from the Mediterranean region and whose essential oil have numerous applications in cosmetics, pharmaceuticals, and alternative medicines. The heterozygous clone "Maillette" was used as a reference for DNA and RNA sequencing. We first built a reference Unigene, compound of coding sequences, thanks to de novo RNA-seq assembly. Then, we reconstructed the complete genes sequences (with introns and exons) using an Unigene-guided DNA-seq assembly approach. This aimed to maximize the possibilities of finding polymorphism between genetically close individuals despite the lack of a reference genome. Finally, we used these resources for SNP mining within a collection of 16 commercial lavender clones and tested the SNP within the scope of a genetic distance analysis. We obtained a cleaned reference of 8, 030 functionally in silico annotated genes. We found 359K polymorphic sites and observed a high SNP frequency (mean of 1 SNP per 90 bp) and a high level of heterozygosity (more than 60% of heterozygous SNP per genotype). On overall, we found similar genetic distances between pairs of clones, which is probably related to the out-crossing nature of the species and the restricted area of cultivation. The proposed method is transferable to other orphan species, requires little bioinformatics resources and can be realized within a year. This is also the first reported large-scale SNP development on Lavandula angustifolia. All the genomics resources developed herein are publicly available and provide a rich pool of molecular resources to explore and exploit lavender genetic diversity in breeding programs.

摘要

下一代测序(NGS)技术通过降低测序成本和提高通量,为以前研究较少的物种生成基因组数据开辟了道路。在这项研究中,我们提出了一种快速开发大规模分子资源的方法,用于研究孤儿物种。我们以多年生亚灌木植物薰衣草(Lavandula angustifolia Mill.)为例,该植物原产于地中海地区,其精油在化妆品、制药和替代医学中具有多种应用。异质克隆“Maillette”被用作 DNA 和 RNA 测序的参考。我们首先通过从头 RNA-seq 组装构建了一个包含编码序列的参考 Unigene。然后,我们使用 Unigene 指导的 DNA-seq 组装方法重建了完整的基因序列(包括内含子和外显子)。这旨在最大限度地利用遗传上接近的个体之间存在多态性的可能性,尽管缺乏参考基因组。最后,我们使用这些资源在 16 个商业薰衣草克隆的集合中进行 SNP 挖掘,并在遗传距离分析的范围内测试 SNP。我们获得了 8030 个功能注释基因的清洁参考序列。我们发现了 359K 个多态性位点,观察到 SNP 频率很高(平均每 90bp 有 1 个 SNP),杂合度水平很高(每个基因型的杂合 SNP 超过 60%)。总的来说,我们发现克隆对之间的遗传距离相似,这可能与该物种的异交性质和有限的种植区域有关。所提出的方法可转移到其他孤儿物种,所需的生物信息学资源很少,并且可以在一年内实现。这也是首次在 Lavandula angustifolia 上报道的大规模 SNP 开发。本文开发的所有基因组学资源均可公开获取,并为探索和利用薰衣草遗传多样性提供了丰富的分子资源,可用于培育计划。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/be15/7732122/b5149246582f/pone.0243853.g001.jpg

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