When sampling an item or surface for DNA, the collection of 'background' DNA (bDNA) from previous use poses an issue as it may impact the detectability of 'target' DNA and the interpretation of the DNA results given alleged activities. This study investigates the prevalence and transferability of bDNA on flooring surfaces within occupied houses under conditions similar to those that are encountered in casework. To assess bDNA presence and transferability, and the impact of how and who contacts the surface, areas used frequently and infrequently were targeted in the kitchen, living room, bedroom and bathroom of five houses, and two samples taken from each area; one directly from the floor and another from a cotton surface after contacting the floor. DNA was detected in 97 % (of 39) of samples collected directly from flooring, with 92 % providing interpretable profiles. DNA was detected in 85 % (of 39) samples collected from cotton swatches after contacting the floors, with 79 % providing interpretable profiles. The overall quantity, number of contributors, and likelihood of observing a major contributor was greater for samples obtained directly from the floor compared to the cotton. In 80 % of samples recovered from cotton, the quantity of DNA recovered was less than 20 % of that which was recovered directly from the floor. Overall, no trend was observed between the level of reported activity by occupants within areas of the same room and the quantity of DNA recovered directly from the flooring, the quantity of DNA transferred to and recovered from the cotton, or the number of contributors in resulting DNA profiles. In contrast, greater quantities of DNA were generally obtained from houses with a greater number of occupants. Profile composition was similar for samples collected from different areas of the same room, irrespective of the level of activity and from where the sample was obtained (i.e. directly from the floor or contacting surface). Occupants were often not detected in DNA profiles collected from rooms they were known to use and could be observed in profiles collected from rooms they reportedly did not use. The findings of this preliminary investigation provide an understanding of the complexities of transfer, persistence, prevalence and recovery of DNA traces in houses occupied by multiple people and highlights the need to consider how and who uses a space, in the investigation of criminal activities where DNA traces are recovered from, or have been in contact with, flooring.