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相关概念视频

Real Time RT-PCR02:57

Real Time RT-PCR

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Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...
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RACE - Rapid Amplification of cDNA Ends02:35

RACE - Rapid Amplification of cDNA Ends

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Rapid Amplification of cDNA Ends, or RACE, is one of the most effective methods to obtain a full-length cDNA from an mRNA sequence between a known internal region to the unknown sequence at the 5’ or 3’ end. The unknown region is cloned in the cDNA by a gene-specific primer that binds the known end, and a hybrid primer that attaches a predefined anchor sequence to the unknown end of the cDNA. The sequence in between is amplified by PCR with an anchor primer and a gene-specific...
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Sanger Sequencing01:57

Sanger Sequencing

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DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
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Next-generation Sequencing03:00

Next-generation Sequencing

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The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
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RNA-seq03:21

RNA-seq

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RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
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Ribosome Profiling02:24

Ribosome Profiling

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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
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相关实验视频

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Targeted RNA Sequencing Assay to Characterize Gene Expression and Genomic Alterations
11:52

Targeted RNA Sequencing Assay to Characterize Gene Expression and Genomic Alterations

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直接的RNA测序是指直接的RNA测序.

Fatih Ozsolak1, Adam R Platt, Dan R Jones

  • 1Helicos BioSciences Corporation, One Kendall Square, Cambridge, Massachusetts 02139, USA. fatihozsolak@gmail.com

Nature
|September 25, 2009
PubMed
概括
此摘要是机器生成的。

直接RNA测序绕过补充DNA (cDNA) 转换,减少偏差并改善转录分析. 这种创新方法使得高通量,低成本的直接RNA测序能够全面了解转录组.

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科学领域:

  • 基因组学和转录基因组学
  • 分子生物学分子生物学
  • 生物信息学是一种生物信息学.

背景情况:

  • 了解基因组和转录组对于人类生物学和疾病研究至关重要.
  • 目前的转录组分析依赖于互补DNA (cDNA) 转换,引入偏差并限制对降解或少量RNA的分析.
  • 现有的方法阻碍了对全基因组转录和RNA的动态状态的全面理解.

研究的目的:

  • 开发和应用一种直接的单分子RNA测序方法,没有先前的RNA转化为cDNA.
  • 克服当前转录组分析方法的局限性,包括偏差和适合退化的RNA.
  • 为了实现高吞吐量,低成本和无偏差的直接RNA测序,以获得完整的转录组理解.

主要方法:

  • 在不将RNA转换为cDNA的情况下进行了直接单分子RNA测序.
  • 测序了女性分子数量的多基化[多基基基基基基基基基]Saccharomyces cerevisiae RNA.
  • 通过使用涂有多DT寡核酸的表面捕获RNA,并通过天然多A尾部通过合成开始测序.

主要成果:

  • 在没有cDNA转换的情况下直接成功测序RNA,证明了一种新的方法.
  • 观察到转录3'端异质性,为RNA处理提供了洞察力.
  • 识别了多基化小核RNA,扩大了对非编码RNA群体的知识.

结论:

  • 直接RNA测序为高通量和成本效益的转录组分析提供了一条途径.
  • 这种方法最大限度地减少了与cDNA合成相关的偏差,使得更准确的转录表征和量化.
  • 实现了对转录组的全面和无偏见的理解的目标,这对生物和疾病研究至关重要.