SMORE-Seq
Simultaneous Mapping of RNA Ends with Sequencing
SMORE-Seq is an RNA-Seq method to identify TSS and polyadenylation sites (PAS) by sequencing the 5′ and 3′ ends simultaneously (Park et al., 2014). Poly(A)+ RNA is first isolated from total RNA, and the 5′ caps are removed with TAP. The de-capped 5′ ends of the mRNA are ligated to P5 adapters. Next, the mRNA is fragmented before ligating P7 adapters to the 3′ ends of the fragments. The mRNA fragments are reverse-transcribed, PCR amplified, and size-selected to ~250 bp fragments. The selected fragments are PCR-amplified and ready for sequencing.
Advantages:
- Identifies both TSS and PAS from the same RNA library dataset
- More accurate in identifying TSS regions than RNA-Seq due to 5′ de-capping with TAP
- Mapping PAS using degradation intermediates is possible because a phosphatase treatment before TAP is omitted
Disadvantages:
- Full-length mRNA samples are preferred, which can be rare in highly degraded samples
Reagents:
Illumina Library prep and Array Kit Selector
Reviews:
Bagchi D. N. and Iyer V. R. The Determinants of Directionality in Transcriptional Initiation. Trends Genet. 2016;32:322-333
References:
Park D., Morris A. R., Battenhouse A. and Iyer V. R. Simultaneous mapping of transcript ends at single-nucleotide resolution and identification of widespread promoter-associated non-coding RNA governed by TATA elements. Nucleic Acids Res. 2014;42:3736-3749