Aegilops biuncialis 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 biuncialis). Raw sequencing reads are deposited here. Assemblies and annotations will be included once available.

核质之间以及线粒体与质体等细胞质器官之间的恰当相互作用对真核细胞功能至关重要。为了深入探究器官质基因组及核质相互作用在植物发育与胁迫反应中的作用，我们的首要目标是调查小麦及其广义的Triticum-Aegilops复合体中器官质基因组的多样性。继此之后，我们将致力于评估基因组在发育各个阶段以及非生物和生物胁迫反应过程中的动态变化。此项研究的结果对于提升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高效方法，且无需进行全基因组扩增，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中国春小麦（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型重复序列（分别为长度达到10kb和20kb）。先前的研究已经执行了全基因组扩增，并通过手动拼接连续片段以强制形成器官质基因组的单一主环配置，而这可能或不一定反映了小麦器官质基因组的真实原生状态。为了解决长重复序列，并在无需全基因组扩增和手动拼接连续片段的情况下进行从头组装，我们利用低输入PacBio 20kb文库制备方法生成长测序读数。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个多样化的野生种、T. durum、T. aestivum cv. Chinese Spring以及三个小麦异源多倍体系。此外，我们还为许多额外的栽培品种、野生种和异源多倍体系生成了Illumina短读序列。本项目包括其中一个样本（Aegilops biuncialis）的数据。原始测序读数已存档于此。一旦可用，将包括组装和注释信息。