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# nascent
Edmund Miller / <img src="https://openmoji.org/data/color/svg/E040.svg" width=50> @e_miller88 / <img src="https://openmoji.org/data/color/svg/E045.svg" width=50> @emiller88
<a href="https://utdallas.edu"><img src="https://gist.githubusercontent.com/Emiller88/b2247c2883c3209950bebb89d5cb9a5f/raw/5455a28a6b27ffe90f32a947986eb80ebe63c6fa/utdmonogram.svg" width="80%"><img></a>
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# Overview
- Background on Nascent Transcript Identification
- History of Development
- Pipeline Overview
---
#### Background on Nascent Transcript Identification
- Goal is to identify changes in transcription
- Rather than RNAseq which isolates all of the RNA in cell
- Pulling out the transcriptionally activate sites **through metabolical labelling**
- Covering a lot of different assays
- Slight variation in computational pipeline can result in **25% change in results**
---
# Enhancers
- Cis-acting DNA sequences that can increase the transcription of genes
- The human genome contains ++hundreds of thousands++ of enhancers
- Evidence of Enhancer-Promoter interaction from cross-linking assays (3c)
- enhancer RNAs (**eRNA**) have a short half-life so in low abundance
<!-- [^3]: Pennacchio 2013 -->
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# Enhancer-promoter Looping
<!-- Need an image that's simple but shows the eRNA coming from the region -->
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# What the Reads Look Like
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## Nascent Transcript(NT) and Transcription Start Sites(TSS)
- Currently 13+ assays for nascent transcript identification
- "minor changes in sample processing could lead to changes of up to >20% in the final results"
---
# History of Development
- Version 1.0 was developed by Ignacio Tripodi and Margaret Gruca and released Apr 16, 2019
- In 2017, THK lab started working to reproduce Kim et al. 2018 in a second dataset
- Struggled with building a reproducible pipeline in snakemake and creating CI/CD workflows and templates in January 2020
- Found nf-core in March 2020
---
# How far we've come
[Original 2018 presentation](https://github.com/Emiller88/edmundmiller.dev/tree/master/static/presentations/2018-04-25)
---
# Obligatory Metro Map
---
# CHM13 Support
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# Transcript Identification
- GroHMM - Predicts transcripts from aligned GROSeq data in the form of bed files.
- HOMER - Transcript identification from GROSeq data
- PINTS - Identifies transcriptional regulatory elements (TREs) identified from nascent-transcript sequencing.
---
## GroHMM
- R package released in 2015 by Minho Chae, Charles Danko, & Lee Kraus
- Draw back is it requires tuning and is quite memory hungry
- [Recommended we use tunits instead](https://github.com/dankoc/groHMM/issues/4)
---
# HOMER
- Released in 2010 from the Glass lab and continued by Chris Brenner
- Created in a land before docker `configureHomer.pl`
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# PINTS Identification
<!-- TODO Check to see if groseq is giving a full transcript or just a start site -->
---
# PINTS
- Released in 2022 by Yu Lab and Lis Lab
- Determines the TSS site(opposed to entire transcription units)
- PINTS achieves optimal balance among resolution, robustness, sensitivity, specificity and computational resources required.
- Supports 10 assays out of the box
---
## Need help?
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Repository: [`nf-core/nascent`](https://github.com/nf-core/nascent)
Tutorial: [`https://nf-co.re/nascent/2.0.0/usage`](https://nf-co.re/nascent/2.0.0/usage)
Chat: [`https://nf-co.re/join`](https://nf-co.re/join) <img src="https://cdn.brandfolder.io/5H442O3W/at/pl546j-7le8zk-6gwiyo/Slack_Mark.svg" width=7.5%></img>`#nascent`
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---
# References
> Yao, L., Liang, J., Ozer, A. et al. A comparison of experimental assays and analytical methods for genome-wide identification of active enhancers.
> Pennacchio LA, Bickmore W, Dean A, Nobrega MA, Bejerano G. Enhancers: five essential questions. Nat Rev Genet. 2013 Apr;14(4):288-95. doi: 10.1038/nrg3458. PMID: 23503198; PMCID: PMC4445073.
> Chae M, Danko CG, Kraus WL (2015). “groHMM: a computational tool for identifying unannotated and cell type-specific transcription units from global run-on sequencing data.” BMC Bioinformatics, 16(222).
> Heinz S, Benner C, Spann N, et al. Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell. 2010;38(4):576-589. doi:10.1016/j.molcel.2010.05.004
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