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Keywords:

  • chromatin immunoprecipitation;
  • gene expression profiling;
  • gene expression regulation;
  • genomics;
  • sequence analysis

ABSTRACT

Next generation sequencing (NGS) has pushed back the limitations of prior sequencing technologies to advance genomic knowledge infinitely by allowing cost-effective, rapid sequencing to become a reality. Genome-wide transcriptional profiling can be achieved using NGS with either Tag-Seq, in which short tags of cDNA represent a gene, or RNA-Seq, in which the entire transcriptome is sequenced. Furthermore, the level and diversity of miRNA within different tissues or cell types can be monitored by specifically sequencing small RNA. The biological mechanisms underlying differential gene regulation can also be explored by coupling chromatin immunoprecipitation with NGS (ChIP-Seq). Using this methodology genome-wide binding sites for transcription factors, RNAP II, epigenetic modifiers and the distribution of modified histones can be assessed. The superior, high-resolution data generated by adopting this sequencing technology allows researchers to distinguish the precise genomic location bound by a protein and correlate this with observed gene expression patterns. Additional methods have also been established to examine other factors influencing gene regulation such as DNA methylation or chromatin conformation on a genome-wide scale. Within any research setting, these techniques can provide relevant data and answer numerous questions about gene expression and regulation. The advances made by pairing NGS with strategic experimental protocols will continue to impact the research community.