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Cryptic contamination and phylogenetic nonsense.

Linderholm A, Malmström H, Lidén K, Holmlund G, Götherström A - PLoS ONE (2008)

Bottom Line: If it does, the results are rejected as contamination, while if it does not, they are often considered authentic.We show here that human contamination in ancient material may well deviate from local allele frequencies or the distributions to be found among the laboratory workers and archaeologists.We conclude that it is not reliable to authenticate ancient human DNA solely by showing that it is different from what would be expected from people who have handled the material.

View Article: PubMed Central - PubMed

Affiliation: Archaeological Research laboratory, Stockholm University, Stockholm, Sweden. anna.linderholm@arklab.su.se

ABSTRACT
Ancient human DNA has been treated cautiously ever since the problems related to this type of material were exposed in the early 1990s, but as sequential genetic data from ancient specimens have been key components in several evolutionary and ecological studies, interest in ancient human DNA is on the increase again. It is especially tempting to approach archaeological and anthropological questions through this type of material, but DNA from ancient human tissue is notoriously complicated to work with due to the risk of contamination with modern human DNA. Various ways of authenticating results based on ancient human DNA have been developed to circumvent the problems. One commonly used method is to predict what the contamination is expected to look like and then test whether the ancient human DNA fulfils this prediction. If it does, the results are rejected as contamination, while if it does not, they are often considered authentic. We show here that human contamination in ancient material may well deviate from local allele frequencies or the distributions to be found among the laboratory workers and archaeologists. We conclude that it is not reliable to authenticate ancient human DNA solely by showing that it is different from what would be expected from people who have handled the material.

Show MeSH
Presence of an 80bp mtDNA fragment in the 74 screened samples, the seven selected samples and 29 non-human samples.The subset of humans was selected from a quantitative pre-screening. Samples yielding sufficient DNA and containing sufficient bone material for repeated re-extraction, and originating from more than one collection were selected for further processing. Note that the same extracts that was used for the mitochondrial pre-screening was used for the -13910 typing in Linköping, while new extracts were produced in Stockholm/Uppsala. A = Human mitochondrial content in all human samples pre-screened for mitochondrial DNA, B = Human mitochondrial content in selected human samples and C = Human mitochondrial content in non-human samples pre-screened for human mitochondrial DNA.
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pone-0002316-g002: Presence of an 80bp mtDNA fragment in the 74 screened samples, the seven selected samples and 29 non-human samples.The subset of humans was selected from a quantitative pre-screening. Samples yielding sufficient DNA and containing sufficient bone material for repeated re-extraction, and originating from more than one collection were selected for further processing. Note that the same extracts that was used for the mitochondrial pre-screening was used for the -13910 typing in Linköping, while new extracts were produced in Stockholm/Uppsala. A = Human mitochondrial content in all human samples pre-screened for mitochondrial DNA, B = Human mitochondrial content in selected human samples and C = Human mitochondrial content in non-human samples pre-screened for human mitochondrial DNA.

Mentions: We selected seven of the 74 human samples that had yielded significantly more human mitochondrial DNA (>1264 copies/5 µl extract) than the non-human samples (<197 copies/5 µl extract) (Fig. 2, Tab. 1), so that these human samples contained on average 198 times more human copies of the mitochondrial fragment tested for (3951±1296 SE) than the 29 non-human specimens (20±7.4 SE). The human samples were then decontaminated and extracted in two laboratories, Linköping (two independent extractions per sample) and Stockholm (a single extraction per sample), the separate extractions being carried out as independent replications performed by different people in each laboratory. Various negative controls were processed in parallel (a minimum of 18 and 4 non-human specimens and 16 and 3 extraction blanks in the Linköping and Stockholm laboratories respectively). DNA amplification was further performed in three laboratories, Linköping, Stockholm and Uppsala. The samples extracted in the Linköping laboratory were also amplified there, while the samples extracted in Stockholm were divided into two aliquots and further processed in Stockholm and Uppsala. All three laboratories are ancient DNA laboratories properly designed according to previously described standards [3], [10], with airlocks, positive airflow and ceiling UV radiation at night. All three laboratories are located in areas separated from any work with high quality DNA or post-PCR procedures, and the laboratory workers wore full zip suites, facemasks and several layers of gloves when in the laboratories.


Cryptic contamination and phylogenetic nonsense.

Linderholm A, Malmström H, Lidén K, Holmlund G, Götherström A - PLoS ONE (2008)

Presence of an 80bp mtDNA fragment in the 74 screened samples, the seven selected samples and 29 non-human samples.The subset of humans was selected from a quantitative pre-screening. Samples yielding sufficient DNA and containing sufficient bone material for repeated re-extraction, and originating from more than one collection were selected for further processing. Note that the same extracts that was used for the mitochondrial pre-screening was used for the -13910 typing in Linköping, while new extracts were produced in Stockholm/Uppsala. A = Human mitochondrial content in all human samples pre-screened for mitochondrial DNA, B = Human mitochondrial content in selected human samples and C = Human mitochondrial content in non-human samples pre-screened for human mitochondrial DNA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2384008&req=5

pone-0002316-g002: Presence of an 80bp mtDNA fragment in the 74 screened samples, the seven selected samples and 29 non-human samples.The subset of humans was selected from a quantitative pre-screening. Samples yielding sufficient DNA and containing sufficient bone material for repeated re-extraction, and originating from more than one collection were selected for further processing. Note that the same extracts that was used for the mitochondrial pre-screening was used for the -13910 typing in Linköping, while new extracts were produced in Stockholm/Uppsala. A = Human mitochondrial content in all human samples pre-screened for mitochondrial DNA, B = Human mitochondrial content in selected human samples and C = Human mitochondrial content in non-human samples pre-screened for human mitochondrial DNA.
Mentions: We selected seven of the 74 human samples that had yielded significantly more human mitochondrial DNA (>1264 copies/5 µl extract) than the non-human samples (<197 copies/5 µl extract) (Fig. 2, Tab. 1), so that these human samples contained on average 198 times more human copies of the mitochondrial fragment tested for (3951±1296 SE) than the 29 non-human specimens (20±7.4 SE). The human samples were then decontaminated and extracted in two laboratories, Linköping (two independent extractions per sample) and Stockholm (a single extraction per sample), the separate extractions being carried out as independent replications performed by different people in each laboratory. Various negative controls were processed in parallel (a minimum of 18 and 4 non-human specimens and 16 and 3 extraction blanks in the Linköping and Stockholm laboratories respectively). DNA amplification was further performed in three laboratories, Linköping, Stockholm and Uppsala. The samples extracted in the Linköping laboratory were also amplified there, while the samples extracted in Stockholm were divided into two aliquots and further processed in Stockholm and Uppsala. All three laboratories are ancient DNA laboratories properly designed according to previously described standards [3], [10], with airlocks, positive airflow and ceiling UV radiation at night. All three laboratories are located in areas separated from any work with high quality DNA or post-PCR procedures, and the laboratory workers wore full zip suites, facemasks and several layers of gloves when in the laboratories.

Bottom Line: If it does, the results are rejected as contamination, while if it does not, they are often considered authentic.We show here that human contamination in ancient material may well deviate from local allele frequencies or the distributions to be found among the laboratory workers and archaeologists.We conclude that it is not reliable to authenticate ancient human DNA solely by showing that it is different from what would be expected from people who have handled the material.

View Article: PubMed Central - PubMed

Affiliation: Archaeological Research laboratory, Stockholm University, Stockholm, Sweden. anna.linderholm@arklab.su.se

ABSTRACT
Ancient human DNA has been treated cautiously ever since the problems related to this type of material were exposed in the early 1990s, but as sequential genetic data from ancient specimens have been key components in several evolutionary and ecological studies, interest in ancient human DNA is on the increase again. It is especially tempting to approach archaeological and anthropological questions through this type of material, but DNA from ancient human tissue is notoriously complicated to work with due to the risk of contamination with modern human DNA. Various ways of authenticating results based on ancient human DNA have been developed to circumvent the problems. One commonly used method is to predict what the contamination is expected to look like and then test whether the ancient human DNA fulfils this prediction. If it does, the results are rejected as contamination, while if it does not, they are often considered authentic. We show here that human contamination in ancient material may well deviate from local allele frequencies or the distributions to be found among the laboratory workers and archaeologists. We conclude that it is not reliable to authenticate ancient human DNA solely by showing that it is different from what would be expected from people who have handled the material.

Show MeSH