scholarly journals The influence of sample quantity and lysis parameters on the success of ancient DNA extraction from skeletal remains

BioTechniques ◽  
2021 ◽  
Author(s):  
Alina Euskirchen ◽  
Larissa Hartmann ◽  
Janine Mazanec ◽  
Patrick Wittmeier ◽  
Susanne Hummel
Keyword(s):  
Mitochondrial Dna ◽  
Sample Preparation ◽  
Dna Extraction ◽  
Ancient Dna ◽  
Skeletal Remains ◽  
Sample Material ◽  
Chromosomal Dna ◽  
High Quality ◽  
Dna Analyses ◽  
Lysis Time

DNA extraction is of utmost importance in archaeobiology, as it determines the success of further DNA analyses. This study concentrates on the success of ancient DNA extraction using silica spin columns and PCR-based analysis from archaeological skeletal material and investigates the influence of sample quantity, lysis time and lysis temperature during sample preparation. The results show that lysis times ranging from 2 to 48 h are suitable, and that lysis should be carried out at a constant temperature of 56°C. Concerning sample quantity, 10 mg for mitochondrial DNA and 50 mg for chromosomal DNA are sufficient for high quality analyses. Thus invaluable sample material can be saved, and time of sample preparation can be reduced considerably.

scholarly journals Comparison of two ancient DNA extraction protocols for skeletal remains from tropical environments

2018 ◽  
Vol 166 (4) ◽  
pp. 824-836 ◽  
Author(s):  
Maria A. Nieves-Colón ◽  
Andrew T. Ozga ◽  
William J. Pestle ◽  
Andrea Cucina ◽  
Vera Tiesler ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Ancient Dna ◽  
Skeletal Remains ◽  
Tropical Environments

scholarly journals Comparison of two ancient DNA extraction protocols for skeletal remains from tropical environments

2017 ◽  
Author(s):  
Maria A. Nieves-Colón ◽  
Andrew T. Ozga ◽  
William J. Pestle ◽  
Andrea Cucina ◽  
Vera Tiesler ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Ancient Dna ◽  
Extraction Methods ◽  
Optimal Choice ◽  
Skeletal Remains ◽  
Tissue Type ◽  
Dna Fragments ◽  
Illumina Hiseq ◽  
Tropical Environments ◽  
Significant Difference

ABSTRACTObjectivesThe tropics harbor a large part of the world’s biodiversity and have a long history of human habitation. However, paleogenomics research in these climates has been constrained so far by poor ancient DNA yields. Here we compare the performance of two DNA extraction methods on ancient samples of teeth and petrous portions excavated from tropical and semitropical sites in Tanzania, Mexico, and Puerto Rico (N=12).Materials and MethodsAll samples were extracted twice, built into double-stranded sequencing libraries, and shotgun sequenced on the Illumina HiSeq 2500. The first extraction protocol, Method D, was previously designed for recovery of ultrashort DNA fragments from skeletal remains. The second, Method H, modifies the first by adding an initial EDTA wash and an extended digestion and decalcification step.ResultsNo significant difference was found in overall ancient DNA yields or post-mortem damage patterns recovered from samples extracted with either method, irrespective of tissue type. However, Method H samples had higher endogenous content and more mapped reads after quality-filtering, but also higher clonality. In contrast, samples extracted with Method D had shorter average DNA fragments.DiscussionBoth methods successfully recovered endogenous ancient DNA. But, since surviving DNA in ancient or historic remains from tropical contexts is extremely fragmented, our results suggest that Method D is the optimal choice for working with samples from warm and humid environments. Additional optimization of extraction conditions and further testing of Method H with different types of samples may allow for improvement of this protocol in the future.

scholarly journals Reproducible, portable, and efficient ancient genome reconstruction with nf-core/eager

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e10947
Author(s):  
James A. Fellows Yates ◽  
Thiseas C. Lamnidis ◽  
Maxime Borry ◽  
Aida Andrades Valtueña ◽  
Zandra Fagernäs ◽  
...  
Keyword(s):  
Ancient Dna ◽  
Genomic Data ◽  
Extended Version ◽  
Research Groups ◽  
High Quality ◽  
Diverse Range ◽  
Dna Analyses ◽  
Heterogeneous Landscape ◽  
Analytical Standards

The broadening utilisation of ancient DNA to address archaeological, palaeontological, and biological questions is resulting in a rising diversity in the size of laboratories and scale of analyses being performed. In the context of this heterogeneous landscape, we present an advanced, and entirely redesigned and extended version of the EAGER pipeline for the analysis of ancient genomic data. This Nextflow pipeline aims to address three main themes: accessibility and adaptability to different computing configurations, reproducibility to ensure robust analytical standards, and updating the pipeline to the latest routine ancient genomic practices. The new version of EAGER has been developed within the nf-core initiative to ensure high-quality software development and maintenance support; contributing to a long-term life-cycle for the pipeline. nf-core/eager will assist in ensuring that a wider range of ancient DNA analyses can be applied by a diverse range of research groups and fields.

scholarly journals Ancient DNA Methods Improve Forensic DNA Profiling of Korean War and World War II Unknowns

Genes ◽  
2022 ◽  
Vol 13 (1) ◽  
pp. 129
Author(s):  
Elena I. Zavala ◽  
Jacqueline Tyler Thomas ◽  
Kimberly Sturk-Andreaggi ◽  
Jennifer Daniels-Higginbotham ◽  
Kerriann K. Meyers ◽  
...  
Keyword(s):  
World War Ii ◽  
Dna Extraction ◽  
Ancient Dna ◽  
Korean War ◽  
Skeletal Remains ◽  
Dna Profiling ◽  
Library Preparation ◽  
Forensic Dna ◽  
World War ◽  
Preparation Methods

The integration of massively parallel sequencing (MPS) technology into forensic casework has been of particular benefit to the identification of unknown military service members. However, highly degraded or chemically treated skeletal remains often fail to provide usable DNA profiles, even with sensitive mitochondrial (mt) DNA capture and MPS methods. In parallel, the ancient DNA field has developed workflows specifically for degraded DNA, resulting in the successful recovery of nuclear DNA and mtDNA from skeletal remains as well as sediment over 100,000 years old. In this study we use a set of disinterred skeletal remains from the Korean War and World War II to test if ancient DNA extraction and library preparation methods improve forensic DNA profiling. We identified an ancient DNA extraction protocol that resulted in the recovery of significantly more human mtDNA fragments than protocols previously used in casework. In addition, utilizing single-stranded rather than double-stranded library preparation resulted in increased attainment of reportable mtDNA profiles. This study emphasizes that the combination of ancient DNA extraction and library preparation methods evaluated here increases the success rate of DNA profiling, and likelihood of identifying historical remains.

scholarly journals Optimized Protocol for Chelex-based Extraction of DNA from Historical Skeletal Remains and Forensic Trace Samples

2021 ◽  
Author(s):  
Wera M Schmerer
Keyword(s):  
Dna Extraction ◽  
Critical Factor ◽  
Skeletal Remains ◽  
Sample Material ◽  
Soil Environment ◽  
Chelex 100 ◽  
Optimized Protocol ◽  
Method Paper ◽  
Forensic Sample

Abstract PCR-based analysis of skeletonized human remains is a common aspect in both forensic human identification as well as Ancient DNA research. In this, both areas not merely utilize very similar methodology, but also share the same problems regarding quantity and quality of recovered DNA and presence of inhibitory substances in samples from excavated remains. To enable amplification based analysis of the remains, development of optimized DNA extraction procedures is thus a critical factor in both areas.The method paper here presents an optimized protocol for DNA extraction from ancient skeletonized remains using Chelex-100, with improved effectively in yielding amplifiable extracts from sample material excavated after centuries in a soil environment, which consequently have high inhibitor content and overall limited DNA preservation. Further studies showed that the optimized protocol can likewise be utilized for extraction of DNA from common and trace Forensic sample material.

scholarly journals Nondestructive DNA extraction method for mitochondrial DNA analyses of museum specimens

BioTechniques ◽  
2004 ◽  
Vol 36 (5) ◽  
pp. 814-821 ◽  
Author(s):  
Nadin Rohland ◽  
Heike Siedel ◽  
Michael Hofreiter
Keyword(s):  
Mitochondrial Dna ◽  
Dna Extraction ◽  
Extraction Method ◽  
Museum Specimens ◽  
Dna Analyses ◽  
Dna Extraction Method

Quantitative defect studies in semiconductor TEM samples prepared by low angle ion milling

1995 ◽  
Vol 53 ◽  
pp. 512-513
Author(s):  
L. Mulestagno ◽  
J.C. Holzer ◽  
P. Fraundorf
Keyword(s):  
Sample Preparation ◽  
Milling Time ◽  
High Density ◽  
Low Density ◽  
Ion Milling ◽  
High Quality ◽  
Variable Angle ◽  
Major Disadvantage ◽  
Induced Defects

Due to the wealth of information, both analytical and structural that can be obtained from it TEM always has been a favorite tool for the analysis of process-induced defects in semiconductor wafers. The only major disadvantage has always been, that the volume under study in the TEM is relatively small, making it difficult to locate low density defects, and sample preparation is a somewhat lengthy procedure. This problem has been somewhat alleviated by the availability of efficient low angle milling.Using a PIPS® variable angle ion -mill, manufactured by Gatan, we have been consistently obtaining planar specimens with a high quality thin area in excess of 5 × 104 μm2 in about half an hour (milling time), which has made it possible to locate defects at lower densities, or, for defects of relatively high density, obtain information which is statistically more significant (table 1).

scholarly journals Evaluation of DNA Extraction Methods Developed for Forensic and Ancient DNA Applications Using Bone Samples of Different Age

Genes ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 146
Author(s):  
Catarina Xavier ◽  
Mayra Eduardoff ◽  
Barbara Bertoglio ◽  
Christina Amory ◽  
Cordula Berger ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Ancient Dna ◽  
Extraction Method ◽  
Fragment Size ◽  
Molecular Genetic ◽  
Extraction Methods ◽  
Degraded Dna ◽  
Size Analysis ◽  
Dna Fragments ◽  
Dna Amount

The efficient extraction of DNA from challenging samples, such as bones, is critical for the success of downstream genotyping analysis in molecular genetic disciplines. Even though the ancient DNA community has developed several protocols targeting small DNA fragments that are typically present in decomposed or old specimens, only recently forensic geneticists have started to adopt those protocols. Here, we compare an ancient DNA extraction protocol (Dabney) with a bone extraction method (Loreille) typically used in forensics. Real-time quantitative PCR and forensically representative typing methods including fragment size analysis and sequencing were used to assess protocol performance. We used four bone samples of different age in replicates to study the effects of both extraction methods. Our results confirm Loreille’s overall increased gain of DNA when enough tissue is available and Dabney’s improved efficiency for retrieving shorter DNA fragments that is beneficial when highly degraded DNA is present. The results suggest that the choice of extraction method needs to be based on available sample, degradation state, and targeted genotyping method. We modified the Dabney protocol by pooling parallel lysates prior to purification to study gain and performance in single tube typing assays and found that up to six parallel lysates lead to an almost linear gain of extracted DNA. These data are promising for further forensic investigations as the adapted Dabney protocol combines increased sensitivity for degraded DNA with necessary total DNA amount for forensic applications.

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