Technical note: Improved ancient DNA purification for PCR using ion‐exchange columns

Kim, Ki-Jeong; Kim, Kyung-Yong; Jeon, Eun-Hee; Togloom, Ariunaa; Cho, Youn-Ock; Lee, Min-Soo; Lkhagvasuren, Gavaachimed; Choi, Jee‐Hye; Tumen, Dashtseveg; Park, Ae Ja; Kim, Keun-Cheol; Park, Ki-Won; Kim, Jae Hyun; Noh, Maengseok; Yoo, Kwon-Jong; Lee, Kwang-Ho

doi:10.1002/ajpa.20782

Cited by 22 publications

(26 citation statements)

References 33 publications

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“…Previous studies reported that it is difficult to amplify ancient DNA because of fragmentation (28,29). However, we could obtain PCR products of more than 400 bp, which provides convincing evidence that improved DNA extraction lead to an increased PCR success rate for this kind of extensive analysis of ancient human populations (16). In addition, we could directly compare the sequencing data of PCR products that were obtained by 2 different technicians, which validated the genetic information from ancient DNA.…”

Section: Discussionsupporting

confidence: 57%

“…These were heavily discolored during early DNA extraction processing from bone samples, which is evidence for poor preservation of human bones (data not shown). We used an ion-exchange column to purify high quality ancient DNA as previously reported (16). This procedure could also remove unknown PCR inhibitors and microbial contamination, which are hurdles to be overcome while handling ancient DNA (19).…”

Section: Extraction Of Ancient Dna and Determination Of Mitochondrialmentioning

confidence: 99%

“…Ancient DNA of high quality was extracted from the ancient bones according to an improved protocol that we have described (16). Briefly, the bones were fragmented using sterile saw blades, and several millimeters of the bone surface were removed using autoclaved sandpaper.…”

Section: Extraction Of Ancient Human Dnamentioning

confidence: 99%

“…Furthermore, 16,769 bp of mitochondrial DNA have been fully sequenced, making SNP information easily accessible (10,11). As mitochondria are transmitted to the next generation only by the maternal gamete during fertilization, we can assume the maternal heritage of analyzed samples.…”

Section: Introductionmentioning

confidence: 99%

“…Recent work suggests that it is indispensable to correctly determine mitochondrial haplogroups by analyzing both coding and control regions of mitochondrial DNA. Our interdisciplinary approach has overcome several obstacles for genetic analysis of human beings from the past (16). We have improved a method for the extraction of high-quality ancient human DNA and strategically amplified several coding and control regions in order to analyze SNPs.…”

Section: Introductionmentioning

confidence: 99%

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Mitochondrial DNA analysis of ancient human bones excavated from Nukdo island, S.Korea

Kim

Kim²,

Choi³

et al. 2010

BMB Reports

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We have performed analyses using ancient DNA extracted from 25 excavated human bones, estimating around the 1 st century B.C. Ancient human bones were obtained from Nukdo Island, which is located off of the Korean peninsula of East Asia. We made concerted efforts to extract ancient DNA of high quality and to obtain reproducible PCR products, as this was a primary consideration for this extensive kind of undertaking. We performed PCR amplifications for several regions of the mitochondrial DNA, and could determine mitochondrial haplogroups for 21 ancient DNA samples. Genetic information from mitochondrial DNA belonged to super-haplogroup M, haplogroup D or its sub-haplogroups (D4 or D4b), which are distinctively found in East Asians, including Koreans or Japanese. The dendrogram and principal component analysis based on haplogroup frequencies revealed that the Nukdo population was close to those of the East Asians and clearly distinguished from populations shown in the other regions. Considering that Nukdo is geologically isolated in the southern part of the Korean peninsula and is a site of commercial importance with neighboring countries, these results may reflect genetic continuity for the habitation and migration of ethnic groups who had lived in a particular area in the past. Therefore, we suggest that phylogenetic analyses of ancient DNA have significant advantages for clarifying the origins and migrations of ethnic groups, or human races. [BMB reports 2010; 43(2): 133-139]

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Section: Discussionsupporting

confidence: 57%

Section: Extraction Of Ancient Dna and Determination Of Mitochondrialmentioning

confidence: 99%

Section: Extraction Of Ancient Human Dnamentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mitochondrial DNA analysis of ancient human bones excavated from Nukdo island, S.Korea

Kim

Kim²,

Choi³

et al. 2010

BMB Reports

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Technical note: Removal of metal ion inhibition encountered during DNA extraction and amplification of copper‐preserved archaeological bone using size exclusion chromatography

Matheson

Marion

Hayter

et al. 2009

American J Phys Anthropol

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A novel technique for the removal of metal ions inhibiting DNA extraction and PCR of archaeological bone extracts is presented using size exclusion chromatography. Two case studies, involving copper inhibition, demonstrate the effective removal of metal ion inhibition. Light microscopy, SEM, elemental analysis, and genetic analysis were used to demonstrate the effective removal of metal ions from samples that previously exhibited molecular inhibition. This research identifies that copper can cause inhibition of DNA polymerase during DNA amplification. The use of size exclusion chromatography as an additional purification step before DNA amplification from degraded bone samples successfully removes metal ions and other inhibitors, for the analysis of archaeological bone. The biochemistry of inhibition is explored through chemical and enzymatic extraction methodology on archaeological material. We demonstrate a simple purification technique that provides a high yield of purified DNA (>95%) that can be used to address most types of inhibition commonly associated with the analysis of degraded archaeological and forensic samples. We present a new opportunity for the molecular analysis of archaeological samples preserved in the presence of metal ions, such as copper, which have previously yielded no DNA results.

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Technical note: Efficiency of total demineralization and ion‐exchange column for DNA extraction from bone

Seo

Zhang

Kim

et al. 2009

American J Phys Anthropol

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We investigated whether a combination of recently introduced methods, total demineralization and ion-exchange columns, would increase DNA recovery from old bone. Ten bone samples taken after a burial period of approximately 60 years were used in this study. Bone powder was digested using total or incomplete demineralization. DNA was extracted by the standard organic method. The DNA extract was purified with ion-exchange columns or QIAquick spin columns. The efficiency of different DNA extraction methods was compared in terms of DNA concentration, inhibitors generated by real-time PCR, and conventional STR typing results. The mean DNA concentration using the total demineralization method is approximately 3 times higher than that using the incomplete demineralization method. For DNA purification, the method using QIAquick spin columns appeared to yield approximately double the DNA than the method using ion-exchange columns. Furthermore, 2 out of 10 samples showed higher levels of inhibition with C(T) values of IPC > or =30 cycles when using only ion-exchange columns. In STR results, total demineralization yielded more locus profiles by 4.2 loci than incomplete demineralization, and QIAquick spin columns also yielded more locus profiles by 3.5 loci than ion-exchange columns. Total demineralization of bone powder significantly increased DNA yield and improved STR typing results. However, the use of ion-exchange columns was not efficient when compared with the method using QIAquick spin columns. It is suggested that the combination of total demineralization and QIAquick spin columns lead to greatly improved STR typing results.

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Technical note: Improved ancient DNA purification for PCR using ion‐exchange columns

Cited by 22 publications

References 33 publications

Mitochondrial DNA analysis of ancient human bones excavated from Nukdo island, S.Korea

Mitochondrial DNA analysis of ancient human bones excavated from Nukdo island, S.Korea

Technical note: Removal of metal ion inhibition encountered during DNA extraction and amplification of copper‐preserved archaeological bone using size exclusion chromatography

Technical note: Efficiency of total demineralization and ion‐exchange column for DNA extraction from bone

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