FourCSeq is an R package dedicated to the analysis of (multiplexed) 4C sequencing data. The package provides a pipeline to detect specific interactions between DNA elements and identify differential interactions between conditions. The statistical analysis in R starts with individual bam files for each sample as inputs. To obtain these files, the package contains a python script (extdata/python/demultiplex.py) to demultiplex libraries and trim off primer sequences. With a standard alignment software the required bam files can be then be generated.