ABR-PG: The Use of Pentaploid Surrogates for Assembly and Anchoring of Octoploid Strawberry Genomes

栽培草莓(Fragaria X Ananassa)及其直系祖先F.Chiloensis和F.virginiana，从进化论、基因组学和园艺学的角度来看，是一个重要的和有趣的作物植物系统。栽培的草莓起源于18世纪中期它的两个直系祖先的杂交，是最年轻的作物物种之一。它有四个基因组，每个基因组有两个副本，总共有八个基因组。这些都是从多达四个祖先那里传下来的，因此栽培的草莓是基因最复杂的农作物之一。到目前为止，还不可能获得栽培草莓的基因组序列，主要是因为有八个基因组的巨大复杂性。在这个项目中，通过实施一项新的战略来应对这一挑战，该战略将需要测序的基因组数量从8个减少到4个。方法是先将一个经过充分研究的两个基因组(称为二倍体)的品种与一个八个基因组的品种杂交，获得一个有五个基因组的杂交品种--称为五倍体。在对五倍体进行测序后，二倍体的基因组被生物信息学减去，只剩下四个基因组需要组装。这一结果将是栽培草莓的直系祖先之一的第一个组装的基因组序列，这将是一个巨大的资源，对于草莓研究人员和育种者来说，他们有兴趣进一步改良这种重要的经济和健康的水果作物。八倍体草莓的参考基因组尚未建立。这些努力面临的挑战包括多倍体固有的挑战，以及与杂合性相关的挑战。八倍体草莓物种，包括栽培的草莓及其近亲F.Chiloensis和F.virginiana，受到近交衰退的影响，用于桃子和苹果基因组计划中基因组组装的加倍单倍体克隆在草莓中并不是一个可用的选择。将实施一种大胆的方法来解决这个问题，将五倍体杂交种用作全基因组测序和连锁作图的“改良单倍体”对象。该项目的目标是构建一个高分辨率的、基于五倍体的连锁图谱，并使用它来锚定基于五倍体的基因组组装，从其中数字减去已知的二倍体基因组拷贝。连锁作图将使用来自Axiom IStraw90 SNP阵列的标记来完成。具体的项目目标是：1)基于新的五倍体作图群体构建高分辨率的八倍体连锁图谱；2)基于新产生的DNA序列和来自单个五倍体植物的光学作图数据构建八倍体基因组组装；3)将基因组组装锚定在连锁图谱上；以及4)比较新组装的八倍体基因组和现有二倍体基因组的效用，作为八倍体种质中SNP发现的扩展计划的参考。该项目将为后续旨在确定八倍体Fragaria物种的亚基因组组成(S)的研究奠定必要的知识基础。只有在这个知识框架内，才有可能确定具有经济和基本科学价值的基因的同源同源和亚基因组特性。利用五倍体后代群体构建连锁图谱将提供对等位基因分离模式的详细洞察，否则这些模式可能会被八倍体基因调用的挑战所掩盖，从而提供区分二体和多体遗传模式的可能性，并最终确定后者是否确实发生在八倍体水平上。待开发的基因组资源，包括第一个八倍体参考基因组，将通过国家生物技术信息中心(http://www.ncbi.nlm.nih.gov/))和蔷薇科基因组数据库(http://www.rosaceae.org/))公共数据库迅速传播。这些研究成果将促进草莓的基础研究和应用研究，并将加强草莓分子标记辅助育种的努力。

The cultivated strawberry (Fragaria x ananassa) and its immediate ancestors, F. chiloensis and F. virginiana, are an important and intriguing crop plant system from evolutionary, genomic and horticultural perspectives. Having originated from a cross between its two immediate ancestors in the mid-1700s, the cultivated strawberry is one of the youngest crop species. It has four genomes, each present in two copies, making a total of eight genomes. These were passed down from as many as four ancestors, thus making the cultivated strawberry among the most genomically complex crop plants. As yet, it has not been possible to get a genome sequence for the cultivated strawberry, primarily because of the great complexity of having eight genomes. This challenge is addressed in this project by implementation of a new strategy that reduces the number of genomes to be sequenced from eight to four. The approach is to get a hybrid with five genomes - called a pentaploid - by first crossing a well-studied variety with two genomes (termed a diploid) to one with eight. After sequencing the pentaploid, the genome of the diploid is subtracted out bioinformatically, leaving only four genomes to be assembled. The result will be the first assembled genome sequence for one of the immediate ancestors of the cultivated strawberry, which will be a resource of immense value to strawberry researchers and breeders interested in further improving this economically important and healthful fruit crop.No reference genome has yet been established for an octoploid strawberry. Challenges facing such efforts include those inherent to polyploidy, as well as those associated with heterozygosity. The octoploid strawberry species, including the cultivated Fragaria x ananassa and its immediate ancestors F. chiloensis and F. virginiana, are subject to inbreeding depression, and doubled haploid clones as used for genome assembly in the peach and apple genome projects have not been an available option in strawberry. A bold approach will be implemented to solve this problem by using pentaploid hybrids as "modified haploid" subjects for whole genome sequencing and linkage mapping. The project goals are to construct a high resolution, pentaploid-based linkage map and use it to anchor a pentaploid-based genome assembly, from which the known diploid genome copy has been digitally subtracted out. Linkage mapping will be accomplished using markers from the Axiom IStraw90 SNP array. The specific project objectives are to: 1) construct a high resolution, octoploid linkage map based upon a novel, pentaploid mapping population; 2) construct an octoploid genome assembly based upon newly generated DNA sequence and optical mapping data derived from a single pentaploid plant; 3) anchor the genome assembly to the linkage map; and, 4) compare the utility of the newly assembled octoploid genome to that of the existing diploid genome as references in relation to an expanded program of SNP discovery in octoploid germplasm. The project will establish a needed knowledge foundation for subsequent research aimed at defining the subgenome composition(s) of the octoploid Fragaria species. Only within this knowledge framework will it be possible to define the homoeologues and subgenomic specificities of genes of economic and basic scientific interest. The employment of a pentaploid progeny population for linkage map construction will provide detailed insight into patterns of allele segregation that might otherwise be obscured by the challenges of genotype calling in an octoploid, thereby offering the potential to differentiate disomic and polysomic inheritance patterns and to finally establish whether the latter do in fact occur at the octoploid level in strawberry. The genomic resources to be developed, including the first octoploid reference genome, will be rapidly disseminated via the National Center for Biotechnology Information (http://www.ncbi.nlm.nih.gov/) and Genome Database for Rosaceae (http://www.rosaceae.org/) public databases. The research products will advance both basic and applied research in strawberry, and will reinforce efforts to implement marker-assisted breeding in strawberry.