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Transcriptomics using next generation sequencing technologies.
Dasfne Lee-Liu1, Leonardo I Almonacid, Fernando Faunes
1Center for Aging and Regeneration and Millennium Nucleus in Regenerative Biology, Pontificia Universidad Catolica de Chile, Santiago, Chile.
Methods in Molecular Biology (Clifton, N.J.)
|September 8, 2012
Summary
This guide introduces RNA sequencing (RNA-Seq) for Xenopus transcriptomics research. It covers essential steps from sample preparation to bioinformatics analysis for gene expression studies.
Area of Science:
- Molecular Biology
- Genomics
- Bioinformatics
Background:
- Next-generation sequencing (NGS) enables transcriptomics studies.
- RNA sequencing (RNA-Seq) provides comprehensive transcriptional profiles.
- Xenopus is a valuable model organism for developmental biology research.
Purpose of the Study:
- To provide a starting point for Xenopus researchers conducting RNA-Seq studies.
- To outline key experimental and computational procedures.
- To guide the interpretation of deep sequencing data in Xenopus.
Main Methods:
- Template isolation and library preparation for RNA-Seq.
- Bioinformatics pipeline for raw data processing and normalization.
- Differential gene expression analysis using RNA-Seq data.
Main Results:
- Detailed methodology for Xenopus RNA-Seq.
- Bioinformatic workflow for transcriptomic data analysis.
- Strategies for interpreting deep sequencing results in Xenopus.
Conclusions:
- RNA-Seq is a powerful tool for Xenopus transcriptomics.
- A standardized approach facilitates robust gene expression studies.
- This chapter serves as a practical resource for Xenopus researchers.
Related Concept Videos
03:00Next-generation Sequencing
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
Although all next-generation methods use different technologies, they all share a set of standard features.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.
03:21RNA-seq
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 microarray-based...
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...
02:02Genomics
Genomics is the science of genomes: it is the study of all the genetic material of an organism. In humans, the genome consists of information carried in 23 pairs of chromosomes in the nucleus, as well as mitochondrial DNA. In genomics, both coding and non-coding DNA is sequenced and analyzed. Genomics allows a better understanding of all living things, their evolution, and their diversity. It has a myriad of uses: for example, to build phylogenetic trees, to improve productivity and...
02:24Ribosome Profiling
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.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
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03:36Genome Annotation and Assembly
The genome refers to all of the genetic material in an organism. It can range from a few million base pairs in microbial cells to several billion base pairs in many eukaryotic organisms. Genome assembly refers to the process of taking the DNA sequencing data and putting it all back together in a correct order to create a close representation of the original genome. This is followed by the identification of functional elements on the newly assembled genome, a process called genome annotation.
02:51Multi-species Conserved Sequences
Next-generation sequencing technologies have created large genomic databases of a variety of animals and plants. Ever since the human genome project was completed, scientists studied the genome of primates, mammals, and other phylogenetically distant living beings. Such large-scale studies have provided new insights into the evolutionary relationship between organisms.
Although the genome of each species varies greatly from each other, a few sequences are highly conserved. Such conserved DNA...
Although the genome of each species varies greatly from each other, a few sequences are highly conserved. Such conserved DNA...