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所在平台: Udemy |
课程主页: https://www.udemy.com/course/next-generation-sequencing-ngs/
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课程名称：下一代测序（NGS） 概述：在过去的56年里，研究人员一直在开发方法和技术，以帮助确定生物样本中的核酸序列。精准测序DNA和RNA对多个研究领域产生了重大影响。人类基因组计划于2003年完成，耗时13年，投入30亿美元，采用了自1970年代以来的主要测序方法——桑格测序。如今，测序需求呈指数级增长，需要快速、廉价且准确地分析大量基因组DNA。下一代测序（NGS）作为新技术的统称，正是应运而生。NGS是一种大规模并行测序技术，具有超高通量、可扩展性和快速性，可以确定整个基因组或DNA和RNA的特定区域的核苷酸顺序。 与桑格测序相比，NGS的主要差异在于测序的体量，NGS可以同时处理数百万个反应，提高了通量和敏感性，同时缩短成本。许多使用桑格测序方法需要多年的基因组测序项目，现在使用NGS可在数小时内完成。NGS的主要步骤包括样本准备（预处理）、文库准备、PCR扩增、测序和数据分析（后处理）。文库准备涉及DNA破碎、末端修复和接头连接；文库扩增确保从测序仪接收到的信号强度足以被准确检测。常见的PCR扩增方法包括乳液PCR和桥接PCR。 多家公司开发了几种竞争性的NGS方法，包括454脉冲测序、配对连接测序（SOLiD）、离子Torrent半导体测序和可逆终端测序（Illumina）。NGS可以生成两种类型的读取，即单端读取和配对端读取，具体取决于所选择的方法。NGS使研究人员能够收集大量的基因组测序数据，具有广泛的应用，包括疫情管理、复杂疾病的诊断与理解、全基因组测序、转录组测序、癌症治疗、病毒检测和抗微生物耐药性监测等。此外，第三代（3G）和第四代（4G）技术也在不断演进。 本课程是与分子生物学、法医科学、医学实验室技术、生物技术和遗传学相关的学生和研究人员的重要资源。现在开始你的学习旅程，探索测序技术背后的隐藏真相吧！
Over the last 56 years, researchers have been developing methods and technologies to assist in the determination of nucleic acid sequences in biological samples. The ability to sequence DNA and RNA accurately has had a great impact in numerous research fields. The sequencing of the human genome was completed in 2003, after 13 years of international collaboration and investment of USD 3 billion. The Human Genome Project used Sanger sequencing, the principal method of DNA sequencing since its invention in the 1970s. Today, the demand for sequencing is growing exponentially, with large amounts of genomic DNA needing to be analyzed quickly, cheaply, and accurately. Thanks to new sequencing technologies known collectively as Next Generation Sequencing.Next generation sequencing (NGS), also known as high throughput sequencing or second generation sequencing or short read sequencing, is a massively parallel sequencing technology that offers ultra-high throughput, scalability, and speed. The technology is used to determine the order of nucleotides in entire genomes or targeted regions of DNA or RNA. The main difference between Sanger sequencing and NGS stems from sequencing volume, with NGS allowing the processing of millions of reactions in parallel, resulting in high-throughput, higher sensitivity, speed and reduced cost. A plethora of genome sequencing projects that took many years with Sanger sequencing methods could now be completed within hours using NGS.There are several main steps that must be tailored to the target (RNA or DNA) and sequencing system selected. The main steps of NGS includes sample preparation (pre-processing), library preparation, PCR amplification, sequencing and data analysis (Post-processing). Library preparation includes DNA fragmentation either enzymatically or by sonication, end repairing and adaptor ligation.Library amplification is required so that the received signal from the sequencer is strong enough to be detected accurately. The two most common PCR amplification methods are emulsion PCR and bridge PCR.Several competing methods of NGS have been developed by different companies including 454 Pyrosequencing, sequencing by ligation (SOLiD), ion torrent semiconductor sequencing and reversible terminator sequencing (illumina).NGS can generate two types of reads i.e, single end reads and paired end reads, depending on method of choice.NGS has enabled researchers to collect vast quantities of genomic sequencing data. This technology has a plethora of applications, such as: outbreak Management, diagnosing and understanding complex diseases, whole-genome sequencing, transcriptome sequencing, cancer treatments, detection of viruses, surveillance of antimicrobial resistance and many more.Now, third (3G) and fourth (4G) generation technologies have been evolved that work on different underlying principles.This course is a valuable resource for students and researchers related to molecular biology, forensic science, medical laboratory technology, biotechnology, and genetics. Start your learning journey now and explore the hidden truth about sequencing technology!
