High-Throughput Genome-Wide Translocation Sequencing HTGTS was developed to study translocation mechanisms in mammalian cells (Chiarle et al., 2011). This approach could be applied for studying antibody repertoires (HTGTS-Rep-seq) (Lin et al., 2016) and CRISPR/Cas9 genome modifications (Mei et al., 2016). In HTGTS-Rep-seq, genomic DNA from […]

Genome-Wide, Unbiased Identification of DSBs Enabled by Sequencing GUIDE-seq (Tsai et al., 2015) relies on the integration of double-stranded oligodeoxynucleotides (DSOs) into DSBs. It belongs to a family of methodsãsuch as HTGTS (Chiarle et al., 2011), LAM-HTGTS (Hu et al., 2016), and Digenome-seq (Kim et […]

Map DNA Double-Strand Breaks DSB-Seq maps in vivo DNA DSBs on a genome-wide scale (Baranello et al., 2014). In this method, DNA is biotinylated by a 3ê-end tailing reaction with biotin-16-dUTP. The biotinylated DNA is sonicated and captured with streptavidin-coated beads. The free DNA fragments […]

Breaks labeling and Enrichment on Streptavidin and Sequencing BLESS is a genome-wide approach to map DSBs at nucleotide resolution (Crosetto et al., 2013). BLESS is able to detect telomere ends, Sce endonuclease_induced DSBs, and complex genome-wide DSB landscapes.In this method, DSBs are ligated in situ […]

Targets of RNA-Binding Proteins Identified by Editing TRIBE identifies the target RNA sequences of RBPs in vivo by modifying the RNA sequence using fusion proteins (McMahon et al., 2016). The fusion protein consists of the RBP of interest, which binds to the target RNA, and […]

Targeted Purification of Polysomal mRNA Targeted purification of polysomal mRNA (TRAP-Seq) maps translating mRNAs under various conditions (Jiao et al., 2010). In this method, tagged ribosomal proteins are expressed in cells. The tagged ribosomal proteins are purified and the RNA isolated. The RNA is reverse-transcribed […]

RNA Immunoprecipitation Sequencing RIP-Seq maps the sites at which proteins are bound to the RNA within RNA-protein complexes (Zhao et al., 2010). In this method, RNA-protein complexes are immunoprecipitated with antibodies targeted to the protein of interest. After RNase digestion, RNA protected by protein binding […]

Ribosome Profiling Sequencing Ribosome profiling (Ribo-Seq), also called active mRNA translation sequencing (ARTseq), isolates RNA that is being processed by the ribosome in order to monitor the translation process (Ingolia et al., 2009). In this method, ribosome-bound RNA first undergoes digestion. The RNA is extracted, […]

RNA Bind-n-Seq RBNS characterizes RBPs by high-throughput quantification of their binding affinity, dissociation constants, and the effects of secondary RNA structures on binding (Lambert et al., 2014). RBNS performs deep sequencing on random short RNA oligonucleotides bound to pools of fluorescently labeled RBPs of different […]

Crosslinking and Immunoprecipitation of RNA Polymerase II Pol II CLIP was developed to isolate and sequence noncoding RNAs involved in transcriptional regulation, particularly RNAPII-mediated transcription (Li et al., 2015). Briefly, cells are UV-crosslinked at 254 nm and lysed. Anti-RNAPII antibodies are used to immunoprecipitate the […]