Abstract:
:High-throughput transcriptional analysis has unveiled a myriad of novel RNAs. However, technical constraints in RNA sequencing library preparation and platform performance hamper the identification of rare transcripts contained within the RNA repertoire. Herein we present targeted-RNA directional sequencing (TARDIS), a hybridization-based method that allows subsets of RNAs contained within the transcriptome to be interrogated independently of transcript length, function, the presence or absence of poly-A tracts, or the mechanism of biogenesis. TARDIS is a modular protocol that is subdivided into four main phases, including the generation of random DNA traps covering the region of interest, purification of input RNA material, DNA trap-based RNA capture, and finally RNA-sequencing library construction. Importantly, coupling RNA capture to strand-specific RNA sequencing enables robust identification and reconstruction of novel transcripts, the definition of sense and antisense RNA pairs and, by the concomitant analysis of long and natural small RNA pools, it allows the user to infer potential precursor-product relations. TARDIS takes ∼10 d to implement.
journal_name
Nat Protocjournal_title
Nature protocolsauthors
Portal MM,Pavet V,Erb C,Gronemeyer Hdoi
10.1038/nprot.2015.120subject
Has Abstractpub_date
2015-12-01 00:00:00pages
1915-38issue
12eissn
1754-2189issn
1750-2799pii
nprot.2015.120journal_volume
10pub_type
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