Bru-Seq
Bru-Seq
Bru-Seq maps nascent RNA transcripts using bromouridine tagging. Active RNAPII synthesizes RNA in the presence of Br-UTP. Tagged RNA transcripts are immunoseparated from total RNA using magnetic beads coated with anti-BrdU antibodies. The captured RNA transcripts are eluted and fragmented before synthesis of cDNA strands via RT and PCR amplification. The resultant cDNA strands are prepared for sequencing using an Illumina TruSeq RNA Library Prep Kit.
Pros:
- Maps sequences of nascent RNA transcripts and determines relative transcription rate
- Detects lncRNAs1
- Detects transcription anywhere on the genome
Cons:
- Limited to cell cultures and other artificial systems, due to the requirement for incubation in the presence of labeled nucleotides
- Bru-Seq: Paulsen M. T., Veloso A., Prasad J., Bedi K., Ljungman E. A., et al. (2013) Coordinated regulation of synthesis and stability of RNA during the acute TNF-induced proinflammatory response. Proc Natl Acad Sci U S A 110: 2240-2245
- 1. Paulsen M. T., Veloso A., Prasad J., Bedi K., Ljungman E. A., et al. (2014) Use of Bru-Seq and BruChase-Seq for genome-wide assessment of the synthesis and stability of RNA. Methods 67: 45-54
- Lefkofsky H. B., Veloso A. and Ljungman M. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells. Mutat Res. 2015;776:9-15
- Kocab A. J., Veloso A., Paulsen M. T., Ljungman M. and Duckett C. S. Effects of physiological and synthetic IAP antagonism on c-IAP-dependent signaling. Oncogene. 2015;