Our Protocols
Protocols from the lab
Analysis of transcription and/or splicing in the nucleus:
Nanopore analysis of co-transcriptional processing (nano-COP):
Drexler HL, Choquet K, Merens HE, Tang PS, Simpson JT, Churchman LS (2021) Revealing nascent RNA processing dynamics with nano-COP. Nature Protocols, 16(3):1343-1375.
scripts for nano-COP data analysis
Updates and corrections to the protocol since publication:
On page 18, when preparing the In-house 10× poly(A) buffer (for poly(A) tailing only), it should read as follows: “Contains 25 mM MnCl2, 10 mM DTT, and 5 mg/ml BSA. Mix 1 μl of 1 M MnCl2, 4 μl of 100 mM DTT, 4 μl of 50 mg/ml BSA, and 31 μl of nuclease-free water.” 1M MnCl2 can be purchased from Sigma (cat. no. M1785).
Subcellular RNA sequencing:
Mayer A, Churchman LS (2017) A Detailed Protocol for Subcellular RNA Sequencing (subRNA-seq). Current protocols in molecular biology / edited by Frederick M. Ausubel... [et al.], 120:4.29.1–4.29.18.
Subcellular RNA-seq protocol PDF
Native elongating transcript sequencing (NET-seq):
In yeast:
Churchman LS, Weissman JS. Native elongating transcript sequencing (NET-seq). Curr Protoc Mol Biol. 2012 Apr;Chapter 4:Unit 4.14.1–17.
scripts for yeast NET-seq analysis
In mammalian cells:
Mayer A, Churchman LS (2016) Genome-wide profiling of RNA polymerase transcription at nucleotide resolution in human cells with native elongating transcript sequencing. Nature Protocols, 11(4):813-33.
updated metazoan protocol:
Martell-Smart D, Ietswaart R, Smalec B, Churchman LS. Profiling metazoan transcription genome-wide with nucleotide resolution using NET-seq (native elongating transcript sequencing) protocols.io. 2020.
scripts for metazoan NET-seq analysis
Analysis of mitochondrial gene expression:
Mitochondrial ribosome profiling (in yeast):
Couvillion MT, Churchman LS (2017) Mitochondrial Ribosome (Mitoribosome) Profiling for Monitoring Mitochondrial Translation In Vivo. Current Protocols in Molecular Biology, 533:4.28.1–4.28.25.