Usage:

mamba.sh [options] <-s species> <-f inputfile.fastq.gz> <-i bowtie2_index_basename> 

Mandatory Arguments:

-f fastq.gz input file a gzip compressed fastq file containing reads from a small RNA-seq experiment reads should be "stranded" (second strand), i.e. the sequence of the read should be the same as the original small RNA. This is the usual configuration of current small RNA-seq protocols

-i bowtie2 index basename of a bowtie2 index for the genome of interest. If your bowtie2 index is /home/user/data/hg38.1.bt2 you should use /home/user/data/hg38 MAmBA assumes that the corresponding fasta is in the same directory with the same basename. If missing, a corresponding fasta file will be created by MAmBA in the output directory and erased at the end of the analysis. If you wish to create a fasta file corresponding to the bowtie2 index in the same folder (so that MAmBA will not spend time to create it) you can run:

bowtie2-inspect /path/to/your/bowtie2index/basename > /path/to/your/bowtie2index/basename.fa

-s species (hsa|mmu) Currently there are only references for human (hg38) and mouse (mm10) which are distributed here: https://sites.google.com/a/uniroma1.it/valeriofulci-eng/software However, MAmBA includes a script (mamba_reference.sh) to generate custom annotation provided that you have annotation files for your favorite species. For details please run:

mamba_reference.sh -h 

-n short_name to use in images MAmBA will output .png images for all pre-miRNAs with sufficient coverage in the output directory. You should provide a human readable name to be used in figures (e.g patient_1 or HeLa_cells). Please avoid spaces in the short name, use underscore.

Command line options:

-o output_directory (default: mamba_out.$infile) MAmBA will create this directory (default: mamba_out.fastq.gz) or, in case it exists already ask for confirmation before overwriting previous output.

-j number_of_threads (default: 1) The number of threads to be used by MAmBA

-a adaptor sequence The sequence of the 3' adapter used in NGS. should be at least 10 nucleotides long only A,C,G,T are allowed. By default is set to "no", will cause adaptor trimming skip, assuming you have already trimmed adapters from your fastq file

-c control spike in The sequence of a control spike-in 5' phosphorylated RNA oligo that you have added to the RNA before bisulfite treatment. If provided, C to U conversion will be estimated (assuming that your RNA oligo only contains unmethylated C). Alternatively, the sequence of an endogenous non-methylated small RNA can be provided (e.g. tRNA(Gly) 5' fragment)

-l log/linear (default: log) If set to any value other than "log" a linear y axis (non-conversion frequency) will be used in figures.

-C color (default: darkgreen) If set the argument will be passed to R as the color to use for bars in figures (useful to plot multiple samples in different colors)

-h|? show this help