A high-throughput platform for assembling genome data: the Saphyr

自2003年人类基因组计划完成以来，经过13年的工作和数十亿美元的投资，产生基因组数据的能力以惊人的速度加速发展。测序技术的进步意味着，仅仅20年后，我们就可以在几个小时内，而不是几年内，以几百美元的成本对一个基因组进行测序。因此，基因组序列数据可以由小型和大型实验室产生，并且许多区域，国家和全球联盟已经新近启动，目标是在未来几年内对全世界数百万个物种进行测序。虽然数以千计的基因组数据集的爆炸式增长创造了新的机会--例如筛选基因组中与抗病性等生态重要性状相关的变异--它也带来了同样严峻的挑战。一个特别的问题涉及基因组数据集的性质;即使在深度测序之后，大多数“基因组”实际上也包含大量不能缝合在一起形成染色体的非连续片段。这种不完全的装配阻碍了连锁、同线性和基因型-表型关联的推断，并且对于以大型复杂基因组为特征的非模式植物、真菌和动物尤其严重。Saphyr平台（Bionano Genomics）通过直接成像兆长荧光条形码光学分子克服了这一障碍。DNA用一种酶处理，这种酶在特定位点沉积荧光标记，然后数百万个大的DNA分子通过专有的纳米通道，它们的图像被转换为图形条形码。使用软件，条形码分子相互比较，并基于荧光标记的匹配模式组装成“光学重叠群”;这些是结构支架，独立于序列，代表非常大的区域。由于这些重叠群很容易跨越序列组装中的断裂，它们允许产生由光学和序列重叠群组成的染色体长度“杂交支架”，其中短读段序列组装体已经过计算机酶处理，使得它们表现出与光学分子相同的条形码模式。光学作图被认为是产生高质量基因组的必要步骤，和许多大型测序联盟使用的管道中的功能，包括新推出的达尔文生命之树项目，该项目旨在捕获约66，000个英国物种的基因组信息。Saphyr是唯一一个高通量的光学图谱平台，但目前还没有专门的设施用于NERC资助的基因组学和环境科学的研究。我们将在QMUL建立一个面向NERC的下一代光学图谱设施，以我们的技术支持为基础，从一系列生物样品和物种中分离超高分子量DNA。这一高通量设施将满足目前生产高质量基因组的需求，从而提高英国在快速发展的环境基因组学领域的技术能力。具体而言，该设施将使研究人员能够（1）在当前和未来的项目中包括基因组组装，（2）在大型基因组网络中发挥更积极的作用，以及（3）利用该平台的多功能性来解决有关特定DNA基序和特征的全基因组分布的新研究问题。在机构层面，Saphyr将使我们能够扩大我们的研究组合，并解决各种非模式生物的生态和进化基因组学的基本问题，这些生物从树木和细菌到哺乳动物和环节动物。此外，Saphyr将加强专业人员的培训，提高我们与其他英国研究机构建立合作的能力，从而成为面向环境的研究界的共同技术资源。

Since the completion of the human genome project in 2003, after 13 years of work and billions of dollars, the ability to produce genomic data has accelerated at phenomenal rates. Advances in sequencing technologies mean that just 20 years on we can now sequence a genome in hours, not years, at the cost of a few hundred dollars. As such, genome sequence data can be produced by small and large labs alike, and numerous regional, national and global consortia have been newly launched with the goal of sequencing millions of species worldwide in the coming years.Unfortunately, while the explosion of thousands of genome datasets has created new opportunities - such as screening genomes for variants that correlate with ecologically-important traits such as disease resistance - it has also created equally steep challenges. A particular problem concerns the nature of genomic datasets; even after deep-sequencing, most "genomes" actually comprise large numbers of non-contiguous fragments that cannot be stitched together into chromosomes. This incomplete assembly obstructs inferences of linkage, synteny, and genotype-phenotype association, and is especially severe for non-model plants, fungi and animals that are characterised by large complex genomes. The Saphyr platform (Bionano Genomics) overcomes this obstacle by directly imaging megabase length fluorescently barcoded optical molecules. DNA is treated with an enzyme that deposits fluorescent marks based at specific sites, and then millions of large DNA molecules are passed through proprietary nanochannels and their images are converted to graphical barcodes. Using software, barcoded molecules are compared against each other, and, based on matching patterns of fluro-marks, assembled into 'optical contigs'; these are structural scaffolds, independent of sequence, that represent very large regions. Since these contigs easily span breaks in sequence assembly, they allow the generation of chromosome length 'hybrid scaffolds' made up of optical and sequence contigs, where the short-read sequence assemblies have been in silico enzyme treated so that they exhibit the same barcode patterns as the optical molecules.Optical mapping is considered an essential step in producing high quality genomes, and features in the pipelines used by many large sequencing consortia, including the newly-launched Darwin Tree of Life Project, which aims to capture the genomic information of the ~66,000 UK species. The Saphyr is the only high-throughput optical mapping platform, yet there is currently no dedicated facility for NERC-funded research working at the interface of genomics and environmental science.We will establish a NERC-oriented next generation optical mapping facility at QMUL, underpinned by our technical support for isolating ultra-high molecular weight DNA from a range of biological samples and species. This high throughout facility will meet current demand for generating high quality genomes, and will thus enhance the UK's technical capacity in the fast developing field of environmental genomics. Specifically, the facility will enable researchers to (1) include genome assemblies in current and future projects, (2) play more active roles in large genome networks, and (3) exploit the platform's versatility to tackle novel research questions regarding the genome-wide distributions of specific DNA motifs and features. At an institutional level, the Saphyr will allow us to expand our research portfolio and address fundamental questions in ecological and evolutionary genomics in diverse non-model organisms, which range from trees and bacteria to mammals and annelids. Moreover, the Saphyr will enhance training of specialised staff, enhance our capacity to establish collaborations with other UK research institutions, and thus emerge as a common technical resource for the environmental-oriented research community.