Aegilops uniaristata organellar-enriched DNA sequencing, assembly, and comparative genomics

Proper interactions between the nucleus and cytoplasmic organelles (mitochondria and plastids) are essential to eukaryotic cellular function. To improve our understanding of the role of organellar genomes and nuclear-cytoplasmic interactions in plant development and stress response, our first aim is to survey organellar genome diversity in wheat and across the broader Triticum-Aegilops complex. This will be followed by work to assess genome dynamics across developmental stages as well as during abiotic and biotic stress response. The results of this work will be important for improving crop traits. To accomplish our goals, it was critical to first establish improved methods for the isolation, sequencing, and assembly of organellar genomes from limited starting material without whole genome amplification. As a proof of concept, we optimized our methods using the Triticum aestivum cv. Chinese Spring, for which there is previous sequencing data available. The mitochondria and chloroplast genomes have large repeats (upto 10kb and 20kb in length, respectively). Previous studies have performed whole genome amplification and have manually stitched contigs to force a single master circle configuration of the organellar genomes, which may or may not reflect the true native state of the wheat organellar genomes. To resolve the long repeats and perform de novo assemblies without whole genome amplification and manual stitching of contigs, we utilized low input PacBio 20kb library preparations to generate long sequencing reads. In total, we sequenced 20 organellar-enriched samples with PacBio, including 13 diverse wild species, T. durum, T. aestivum cv. Chinese Spring, and three wheat alloplasmic lines. In addition we generated Illumina short-read sequences for many additional cultivars, wild species, and alloplasmic lines. This project includes data for one of these samples (Aegilops uniaristata). Raw sequencing reads are deposited here. Assemblies and annotations will be included once available.

核与细胞质器官（如线粒体和叶绿体）之间的适当相互作用对真核细胞功能的实现至关重要。为深化我们对器官质基因组及核质相互作用在植物发育与胁迫反应中作用的认知，我们的首要目标是调查小麦及其更广泛的Triticum-Aegilops复合群中器官质基因组的多样性。此研究将继之以对发育阶段及非生物和生物胁迫反应过程中基因组动态的评估。此项工作的成果将对改良作物性状具有重要意义。为实现我们的目标，首先建立从有限的起始材料中分离、测序和组装器官质基因组的方法，且无需全基因组扩增，显得尤为关键。作为概念验证，我们利用具有先前测序数据的Triticum aestivum cv. Chinese Spring优化了我们的方法。线粒体和叶绿体基因组存在大片段重复（分别为10kb和20kb），先前研究已进行全基因组扩增并手动拼接连续序列以强制构建单一的器官质基因组主环结构，但该结构可能并不反映小麦器官质基因组的真实原生状态。为解决长片段重复并实现无需全基因组扩增和手动拼接连续序列的从头组装，我们采用了低输入PacBio 20kb文库制备方法以生成长测序读数。总计，我们使用PacBio对20个器官质富集样本进行了测序，包括13个多样性野生种、T. durum、T. aestivum cv. Chinese Spring和三个小麦异源染色体线。此外，我们还为许多额外的栽培品种、野生种和异源染色体线生成了Illumina短读序列。本项目包括其中一种样本（Aegilops uniaristata）的数据。原始测序读数已存档于此。一旦组装和注释完成，将包含在内。