Paired-End Genomic Signature Tags: A Method for the Functional Analysis of Genomes and Epigenomes

摘要: Because paired-end genomic signature tags are sequenced-based, they have the potential to become an alternate tool tiled microarray hybridization as a method for genome-wide localization of transcription factors and other sequence-specific DNA binding proteins. As outlined here also can be used global analysis methylation. One advantage this approach is ability easily switch between different genome types without having fabricate new each every type. However, does some disadvantages. Among most rate-limiting steps our PE-GST protocol need concatemerize diTAGs, size fractionate them then clone prior sequencing. This usually followed by additional amplify select long (> or = 500) concatemer inserts These time-consuming important standard sequencing increase efficiency approximately 20-30-fold since amplified now provide information on multiple tags; limitation data acqui- sition read length during development methods such Life Sciences' 454 nanotechnology-based instrument (41) could tag several orders magnitude 100,000 diTAG reads/run), which sufficient in-depth all ChIP PE-GSTs in single run. because lengths diTAGs (approximately 60 bp) fall well within region high accuracy instrument. In principle, sequence begin soon generated, thereby completely bypassing concatemerization, sizing, downstream cloning template purification. addition, places any one unique four-base nucleotide sequences, GATC, pair, help instrument's software keep base register well-located peak height indicator middle feature permit multiplexing simultaneous pooled libraries if linker formation (Figure 4).