Sequencing#RNA sequencing

As RNA sequencing, determining the nucleotide sequence of the RNA is known. For this purpose, the RNA is translated into cDNA, so that the method of DNA sequencing can be used. RNA Seq reveals information for gene expression are expressed as, for example, different alleles of a gene. In the case of RNA -Seq also the recognition of post-transcriptional modifications or identification of fusion genes.

Overview

Basically, you can divide the technologies for the study of gene expression in hybridization- based methods and sequence-based methods. Hybridization -based methods, such as microarrays are relatively cheap, but these methods have some limitations, such as high background noise and lower resolution ( engl. dynamic range ). Sequence -based methods such as Sanger sequencing are very time consuming and expensive, but have evolved into SAGE and RT- PCR.

RNA -Seq is a modern sequence-based method and is based on next generation sequencing ( engl. next- generation sequencing ). RNA -Seq has clear advantages over the other methods. RNA -Seq helps to explore complex transcriptomes and provides information on which come together exons in the messenger RNA. Low background noise, higher resolution and high reproduction rates in technical and biological replicates are clear advantages of RNA -Seq. However, " Next- generation sequencing " techniques are very expensive.

Biological Background

The cell uses only a portion of their genes. These include the housekeeping genes and the genes of specialized cell. For example, muscle cells have mechanical properties and blood cells to transport oxygen. All cells have identical genes, but differ in their gene expression. Gene expression is the synthesis of proteins from DNA. Gene expression analysis or transcriptome analysis measures, which genes are turned on or off. When a gene is turned on, then parts of the gene to be transferred into the mRNA. Methods of gene expression analysis, such as the RNA -Seq, measures the concentration of the mRNA in a variety of experimental conditions (for example with / without drugs). Thus, the gene expression analysis follows the question of how the mRNA concentration behaves by drugs in various stages of development of the cell in a healthy or diseased state.

The RNA sequence can better understand the mechanism of alternative splicing, and fusion genes. Alternative splicing is the process by which the pre- RNA is converted to different mRNAs, and thus in various proteins. Fusion genes are hybrid genes from two previously separate genes, united in a gene. Fusion genes generated by translocation, interstitial deletion, or chromosomal inversion.