Computational protocol: Polysome profiling reveals translational control of gene expression in the human malaria parasite Plasmodium falciparum

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Protocol publication

[…] Coverage plots were prepared by extracting the normalized read counts for the region of interest for all genes included in the analysis, scaling the read counts for each gene (z-score) and subsequently calculating the average value for each nucleotide position. Coverage profiles were smoothed in R using the function smooth.spline with a smoothing parameter of 0.35, and were subsequently plotted using bioconductor R package ggplot2. For the var genes, normalized read counts for exon 1, intron, and exon 2 were extracted separately and were divided into bins of approximately equal length (that is, 650 bins for exon 1, 100 bins for intron and 150 bins for exon 2). The average coverage of each bin was calculated and used for subsequent scaling and averaging across the total length of all var genes. [...] The intron/exon boundaries of all intron-spanning reads (defined by samtools CIGAR string dMdNdM where d is one or more digits) were compared to annotated intron/exon boundaries and were selected for further inspection when one or both intron boundaries were different from current PlasmoDB (version 9.0) annotations. All introns with one unannotated intron/exon boundary supported by at least 5 sequence reads and all novel introns (both intron/exon boundaries unannotated) with at least 10 supporting reads were manually verified in a genome browser and were compared to previously reported alternative splice variants [,,]. […]

Pipeline specifications

Software tools Ggplot2, SAMtools
Applications Miscellaneous, Genome data visualization
Organisms Plasmodium falciparum, Homo sapiens
Diseases Malaria