| Physically isolated pre-PCR-facility with strict decontamination strategy in place using bleach and UV-light, movement only from pre-PCR to post-PCR area |
Use of an aDNA laboratory completely isolated and located in a different building than all post-PCR laboratories (Figure 1); routine application of a strict one way regime (Figure 1.) and decontamination strategies for samples, surfaces, reagents and tools in place |
| Extraction and PCR controls |
Blank controls were performed for each set of extracts or PCR round to a ratio of 1:7 or 1:8 |
| Reproducibility |
Multiple PCR rounds from the same or different extractions of each individual yielded consistent results (Table S1) |
| Quantification of starting templates |
Quantification of the number of starting molecules was carried out by quantitative real time PCR but mainly for methodological reasons and less to prove authenticity |
| Appropriate molecule behaviour |
We observed an inverse correlation between amplification efficiency and length of amplicon (Table 1) |
| UDG treatment (only [16]) |
UDG treatment was carried out prior to each PCR round |
| Cloning |
Direct sequencing of all PCR products was sufficient in this study, cloning was not attempted |
| Independent replication in another aDNA laboratory |
In the present study an independent replication was considered not to be necessary; however, the results of a previous study [9] can be considered as independent replication because the previous study had been carried out in a different laboratory and by different personnel |
| Biochemical preservation |
The biochemical preservation of bone collagen analysed exemplarily for three individuals indicated that at least the specimens of the largest of the three archaeological sites were preserved well enough to allow DNA analysis |
| Preservation of associated remains |
No associated remains e.g. of animals were excavated; instead, human host DNA was amplified, but the significance of the results is limited |
