Next-generation sequencing technologies 下一代测序技术

Next-generation sequencing technologies has improved a lot in 15 years.

Read, the length of one measurable sequence, has been lengthened.Flux, the quantities of data from sequence, has also increased.What is more, the cost of sequencing has been decreased.Though the technology is important, it still has defects.For example, NGS could provide an extremely great quantities of fluxes,but the quality is not accurate enough.Another problem is that large flux and long read cannot be measured at the same time.And I think conquering it is the way from which this technology develops.

Short-read NGS

There are two kinds of methods on short-read NGS, sequencing by ligation(SBL) and sequencing by synthesis(SBS).There are a lot platforms which provide service of NGS.Individual short-read sequencing platforms vary with respect to throughput, cost, error profile and structure.The most common platform is Illumina. Its feature is reliable and stable while its defect is that the method of sequencing is monotonous.Main platforms also include SOLiD system, Complete Genomics system and GeneReader system from Qiagen.

Long-read sequencing approaches fall under two broad categories:

single-molecule real-time sequencing approaches and synthetic approaches that rely on existing short-read technologies to construct long reads in silico.

The instruments that are widely used in the long-read sequencing are PacBio and ONT.

By using PacBio, it is difficult for DNA templates longer than ~ 3kb to be sequenced multiple times, shorter DNA templates can be sequenced many times.

The ONT MinION is a small USB-based device that runs off a personal computer.

This affords the MinION superior portability. But ONE MinION has a large error rate.Fortunately, recent improvements in the chemistry and the base calling algorithms are improving accuracy.

Comparison of short-read sequencing and long-read sequencing.

Short-read sequencing can provide vast quantities of data within a short time, but it has its own limitation.Genomes are highly complex with many long repetitive elements.Many of these complex elements are so long that short-read paired-end technologies are insufficient to resolve them.Long-read sequencing delivers reads in excess of several kilobases, allowing for the resolution of these large structural features.Such long reads can span complex or repetitive regions with a single continuous read, thus eliminating ambiguity in the positions or size of genomic elements.Long reads can also be useful for transcriptomic research, as they are capable of spanning entire mRNA transcripts, allowing researchers to identify the precise comnnectivity of exons and discern gene isoforms. 

To my way of thinking, genome sequencing technology has the capacity for changing human lives, and they will be of great value in curing diseases, improving the ecological environment and so on.Though the existing technology has reached a higher level, there’s an important balance between the sequencing speed and accuracy.Each platform has it own merits, but they each refuse to share their advantages in some ways for their own interests, even combined with political will,.For the sake of technology development and the future of mankind, we should take off the selfishness and exert every effort.

下一代测序技术在15年里进步了很多。Read，一个可测量序列的长度，被延长了。通量，即来自序列的数据量，也增加了。更重要的是，测序的成本已经降低。虽然这项技术很重要，但它仍有缺陷。例如，NGS可以提供大量的通量，但质量不够准确。另一个问题是大通量和长读数不能同时测量。我认为征服它是这项技术发展的方向。

短读NGS有两种方法:连接测序(SBL)和合成测序(SBS)。有很多平台提供NGS服务。单个短读测序平台的吞吐量、成本、错误概况和结构各不相同。最常见的平台是Illumina。其特点是可靠稳定，缺点是测序方法单一。主要平台还包括Qiagen公司的SOLiD系统、Complete Genomics系统和GeneReader系统。长读测序方法分为两大类:单分子实时测序方法和依靠现有短读技术在硅上构建长读的合成方法。在长读测序中广泛使用的仪器是PacBio和ONT。使用PacBio，很难对长度超过3kb的DNA模板进行多次测序，较短的DNA模板可以进行多次测序。

ONT MinION是一种基于usb的小型设备，运行在个人电脑上。

这为MinION提供了优越的可移植性。但是一个奴才有很大的错误率。幸运的是，最近化学和基本调用算法的改进提高了准确性。短读测序和长读测序的比较。短读排序可以在很短的时间内提供大量的数据，但它有自己的局限性。基因组是高度复杂的，有许多长而重复的元素。这些复杂元素中的许多都很长，短读对端技术不足以解决它们。长读测序提供超过几个碱基的读值，允许这些大型结构特征的分辨率。这种长时间的读取可以通过一次连续的读取跨越复杂或重复的区域，从而消除了基因组元素位置或大小的模糊性。长reads也可以用于转录组学研究，因为它们能够跨越整个mRNA转录本，使研究人员能够识别外显子的精确连接和识别基因的亚型。在我看来，基因组测序技术具有改变人类生活的能力，在治疗疾病、改善生态环境等方面具有很大的价值。虽然现有的技术已经达到了更高的水平，但在测序速度和准确性之间有一个重要的平衡。每个平台都有自己的优点，但他们都拒绝在某些方面分享自己的优势，为自己的利益，甚至结合政治意愿。为了科技的发展和人类的未来，我们应该摈弃自私，尽一切努力。