Seqpac provides functions and workflows for analysis of short sequenced reads. It was originally developed for small RNA (sRNA) analysis, but can be implemented on any sequencing raw data (provided as a fastq-file), where the unit of measurement is counts of unique sequences. The core of Seqpac is the generation and subsequent annotation/analysis/ visualization of a standardized object called PAC. Using an target system, Seqpac processes, analyzes and visualizes group differences using the PAC object. Seqpac is particularly useful in generating sequence coverage profiles across reference sequences (such as tRNA or gene coverage) without losing the integrity of the original fastq sequences. Thus, candidate sequences are easily validated by for example a BLAST at a genome browser. This guide will introduce you to the basics of our recommended workflow for sRNA analysis, both fastq-processing (e.g. adapter trimming and generating sequence counts) and post-fastq analysis (e.g filtering, genome and sRNA alignment, differential expression analysis and visualization). To assure package quality, Seqpac has been optimized for Bioconducter.