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The impact of read depth and read length on RNA-seq splicing analysis


This Github repository provides the code the for the publication Ladwig et al. 2026.

Authors

  • Annika Ladwig
  • Melina Klostermann
  • Kathi Zarnack

1 Preprocessing and alternative splicing analysis with MAJIQ


The Rmarkdown contains the code for the preprocessing of the raw reads and for the subsequent alternative splice analysis with MAJIQ.

The rendered HTML file is available at https://annikala.github.io/.

2 Impact of library characteristics on alternative splice analysis


The R scripts contain the code used to investigate the influence of read length (2.1a - U2AF2 KD, 2.1b - UPF1 KD) and read depth (2.2) on:

  • read alignment
  • LSV detection
  • LSV quantification
  • alternative splicing event types distribution

3 Impact of library characteristics on the detection of local splice variations (LSVs) in lowly expressed genes


The R script '3_TPM_analysis.Rmd' contains the code for analysing the impact of read length and read depth on the detection of LSVs in lowly expressed genes.

4 Metaanalysis of public RNA-seq datasets from ENCODE


The R script contains the code for the metaanalysis of public RNA-seq datasets from ENCODE to investigate the used read depth.

5 Metaanalysis of public RNA-seq datasets from SRA


The R script contains the code for the metaanalysis of public RNA-seq datasets from SRA to investigate the used read depth.

6 Simulation of quantifiable introns


The R script contains the code for simulating the number of quantifiable introns depending on read length and read depth. Subsampling of an RNA-seq sample in HEK cells with originally 380 M reads and 150-nt read length is used to estimate the number of introns spanned by at least 10 reads, which we denote as quantifiable introns.

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