Moll Karen M, Zhou Peng, Ramaraj Thiruvarangan, Fajardo Diego, Devitt Nicholas P, Sadowsky Michael J, Stupar Robert M, Tiffin Peter, Miller Jason R, Young Nevin D, Silverstein Kevin A T, Mudge Joann
National Center for Genome Resources, 2935 Rodeo Park Drive East, Santa Fe, NM, 87505, USA.
Montana State University, Center for Biofilm Engineering, Bozeman, MT, 59717, USA.
BMC Genomics. 2017 Aug 4;18(1):578. doi: 10.1186/s12864-017-3971-4.
Third generation sequencing technologies, with sequencing reads in the tens- of kilo-bases, facilitate genome assembly by spanning ambiguous regions and improving continuity. This has been critical for plant genomes, which are difficult to assemble due to high repeat content, gene family expansions, segmental and tandem duplications, and polyploidy. Recently, high-throughput mapping and scaffolding strategies have further improved continuity. Together, these long-range technologies enable quality draft assemblies of complex genomes in a cost-effective and timely manner.
Here, we present high quality genome assemblies of the model legume plant, Medicago truncatula (R108) using PacBio, Dovetail Chicago (hereafter, Dovetail) and BioNano technologies. To test these technologies for plant genome assembly, we generated five assemblies using all possible combinations and ordering of these three technologies in the R108 assembly. While the BioNano and Dovetail joins overlapped, they also showed complementary gains in continuity and join numbers. Both technologies spanned repetitive regions that PacBio alone was unable to bridge. Combining technologies, particularly Dovetail followed by BioNano, resulted in notable improvements compared to Dovetail or BioNano alone. A combination of PacBio, Dovetail, and BioNano was used to generate a high quality draft assembly of R108, a M. truncatula accession widely used in studies of functional genomics. As a test for the usefulness of the resulting genome sequence, the new R108 assembly was used to pinpoint breakpoints and characterize flanking sequence of a previously identified translocation between chromosomes 4 and 8, identifying more than 22.7 Mb of novel sequence not present in the earlier A17 reference assembly.
Adding Dovetail followed by BioNano data yielded complementary improvements in continuity over the original PacBio assembly. This strategy proved efficient and cost-effective for developing a quality draft assembly compared to traditional reference assemblies.
第三代测序技术可产生长达数十千碱基的测序读段，通过跨越模糊区域和提高连续性来促进基因组组装。这对于植物基因组而言至关重要，因为植物基因组由于高重复含量、基因家族扩张、片段和串联重复以及多倍体等原因而难以组装。最近，高通量映射和支架策略进一步提高了连续性。这些长距离技术共同使我们能够以具有成本效益且及时的方式获得复杂基因组的高质量草图组装。
在此，我们展示了使用PacBio、Dovetail Chicago（以下简称Dovetail）和BioNano技术对豆科模式植物蒺藜苜蓿（R108）进行的高质量基因组组装。为了测试这些技术用于植物基因组组装的效果，我们在R108组装中使用这三种技术的所有可能组合和顺序生成了五个组装。虽然BioNano和Dovetail连接重叠，但它们在连续性和连接数量上也显示出互补的提升。两种技术都跨越了PacBio单独无法跨越的重复区域。与单独使用Dovetail或BioNano相比，组合技术，特别是先使用Dovetail再使用BioNano，带来了显著的改进。使用PacBio、Dovetail和BioNano的组合生成了R108的高质量草图组装，R108是在功能基因组学研究中广泛使用的蒺藜苜蓿种质。作为对所得基因组序列实用性的测试，新的R108组装用于确定断点并表征先前鉴定的4号和8号染色体之间易位的侧翼序列，鉴定出先前A17参考组装中不存在的超过22.7 Mb的新序列。
先添加Dovetail数据再添加BioNano数据，在连续性方面相对于原始PacBio组装有互补性的提升。与传统参考组装相比，该策略在开发高质量草图组装方面被证明是高效且具有成本效益的。
