Joint Estimation of Contamination,Error and Demography for Nuclear DNA from Ancient Humans

When sequencing an ancient DNA sample from a hominin fossil, DNA from present-day humans involved in excavation and extraction will be sequenced along with the endogenous material. This type of contamination is problematic for downstream analyses as it will introduce a bias towards the population of the contaminating individual(s). Quantifying the extent of contamination is a crucial step as it allows researchers to account for possible biases that may arise in downstream genetic analyses. Here, we present an MCMC algorithm to co-estimate the contamination rate, sequencing error rate and demographic parameters—including drift times and admixture rates—for an ancient nuclear genome obtained from human remains, when the putative contaminating DNA comes from present-day humans. We assume we have a large panel representing the putative contaminant population (e.g. European, East Asian or African). The method is implemented in a C++ program called ‘Demographic Inference with Contamination and Error’ (DICE). We applied it to simulations and genome data from ancient Neanderthals and modern humans. With reasonable levels of genome sequence coverage (>3X), we find we can recover accurate estimates of all these parameters, even when the contamination rate is as high as 50%.     

Nucleus-Cytosol Interactions--A Source of Stoichiometric Error in Flow Cytometric Estimation of Nuclear DNA Content in Plants

Discrepancies of 2–20% were noted in nuclear DNA contentestimates using mixed and unmixed extracts of several plantspecies combinations (target/standard). This may be becausecytosolic components affect dye accessibility to DNA. Severaldilution experiments showed clearly how cytosol concentrationmodified dye accessibility. Heat treatment suggested that dyeaccessibility could be related to chromatin sensitivity to decondensation.Some methods (nucleus isolation, dilution) can decrease, butnot eliminate, stoichiometric error. The high sensitivity ofthe nucleus to the medium highlights the problem of reliabilitylimits in estimating genome size. The choice between internaland external standardization and the interpretation of intraspecificvariation in ‘nuclear DNA content’ are discussed.Copyright 2000 Annals of Botany Company Propidium iodide, dye accessibility, chromatin condensation, standardization, intraspecific variation     

Logistic方程参数估计中的错误与修正

Logistic方程在种群生态学研究中被广泛应用,其积分式为N=k／（1＋e~（a-rt））,式中e~a=（k-N_0）／N_0。以往对该模型参数的各种估计方法,均将r,K和a作为三个相互独立的参数对待,而与e~a=（K-N_0）/N_0的假设相矛盾。由此估计出的参数K和a使实验初值N_0（t=0时的N值）发生偏离。笔者认为N_0是一个不带随机误差的常数,a值决定于K和N_0,而不是一个独立的参数,因而以往的参数估计方法是错误的,必须予以修正。本文提出了具体的修正方法,即用Marquardt方法或单纯形加速法求参数r和K,然后根据实验初值N_0求a,从而使Logistic微积分方程的共同参数r和K的估计一致。     

Bayesian Estimation of Sequence Damage in Ancient DNA

Mol. Biol. Evol. 24:1416-1422.     

Mitochondrial and Nuclear DNA Survey of Zootoca vivipara across the Eastern Italian Alps: Evolutionary Relationships,Historical Demography and Conservation Implications

The European common lizard Zootoca vivipara exhibits reproductive bimodality, with populations being either viviparous or oviparous. In the central-eastern Italian Alps oviparous populations (Z. v. carniolica) and viviparous populations (Z. v. vivipara) partly overlap geographically. Studying the evolutionary relationship between these taxa presents an interesting opportunity to gain insight into the evolution of this trait. We aim to: i) test whether Z. v. carniolica, which is endangered, constitutes an ESU (Evolutionary Significant Unity); ii) infer mtDNA divergence time between the Z. v. carniolica clade and all the other Z. vivipara subspecies with the aid of an external calibration point; and iii) describe the phylogeographical and demographic scenarios in the area. To do so we sequenced about 200 individuals for mitochondrial variation; 64 of them were also analysed for three nuclear genes. Furthermore, we analysed the same nuclear markers in 17 individuals from the other oviparous subspecies Z. v. louislantzi and 11 individuals of Z. v. vivipara from widespread geographical origins. The mtDNA and nDNA loci that we examined supported the monophyly of Z. v. carniolica. The mtDNA-based estimate of divergence time between Z. v. carniolica and all the other subspecies indicated a separation at 4.5 Mya (95% CI 6.1–2.6), with about 5% of sequence divergence. Considering that Z. v. carniolica harbours higher genetic diversity, while Z. v. vivipara from central-eastern Alps shows a signature of recent population and spatial expansion, we argue that Z. v. carniolica represents a distinct evolutionary unit, with a presumably long-term evolutionary history of separation. Z. v. carniolica populations, occurring at higher latitudes and altitudes than insofar supposed, live in peat bogs, a seriously threatened habitat: taking into account also its evolutionary distinctness, specific conservation measures should be considered.     

Differential Nuclear and Mitochondrial DNA Preservation in Post-Mortem Teeth with Implications for Forensic and Ancient DNA Studies

Major advances in genetic analysis of skeletal remains have been made over the last decade, primarily due to improvements in post-DNA-extraction techniques. Despite this, a key challenge for DNA analysis of skeletal remains is the limited yield of DNA recovered from these poorly preserved samples. Enhanced DNA recovery by improved sampling and extraction techniques would allow further advancements. However, little is known about the post-mortem kinetics of DNA degradation and whether the rate of degradation varies between nuclear and mitochondrial DNA or across different skeletal tissues. This knowledge, along with information regarding ante-mortem DNA distribution within skeletal elements, would inform sampling protocols facilitating development of improved extraction processes. Here we present a combined genetic and histological examination of DNA content and rates of DNA degradation in the different tooth tissues of 150 human molars over short-medium post-mortem intervals. DNA was extracted from coronal dentine, root dentine, cementum and pulp of 114 teeth via a silica column method and the remaining 36 teeth were examined histologically. Real time quantification assays based on two nuclear DNA fragments (67 bp and 156 bp) and one mitochondrial DNA fragment (77 bp) showed nuclear and mitochondrial DNA degraded exponentially, but at different rates, depending on post-mortem interval and soil temperature. In contrast to previous studies, we identified differential survival of nuclear and mtDNA in different tooth tissues. Futhermore histological examination showed pulp and dentine were rapidly affected by loss of structural integrity, and pulp was completely destroyed in a relatively short time period. Conversely, cementum showed little structural change over the same time period. Finally, we confirm that targeted sampling of cementum from teeth buried for up to 16 months can provide a reliable source of nuclear DNA for STR-based genotyping using standard extraction methods, without the need for specialised equipment or large-volume demineralisation steps.     

Estimation of Species Identification Error: Implications for Raptor Migration Counts and Trend Estimation

Abstract: One of the primary assumptions associated with many wildlife and population trend studies is that target species are correctly identified. This assumption may not always be valid, particularly for species similar in appearance to co-occurring species. We examined size overlap and identification error rates among Cooper's (Accipiter cooperii) and sharp-shinned (A. striatus) hawks specific to a raptor migration count station along the Pacific Coast of North America. Illustrating the difficulty of distinguishing between these 2 species, we found overlap in 7 metrics among species-sex groups and in 2 metrics between species, and a principal components analysis revealed a continuum of discrete clusters for each species-sex combination in morphospace. Among juvenile hawks (n = 940), we found the greatest misidentification rate for male Cooper's hawks (23% of the 156 males were identified as sharp-shinned), lesser error rates for female Cooper's (8%, n = 339) and female sharp-shinned (6%, n = 246), and the lowest misidentification rate for male sharp-shinned hawks (0%, n = 199). We observed a similar pattern of misidentification among adult hawks (n = 48). We attempted to use conditional probabilities (identification rates) from calibration data to calculate the true number of adult and juvenile Cooper's hawks and sharp-shinned hawks. Discrepancies between total number of observed accipiters and estimated number using calibration data suggest that daily observer misclassification rates are higher than misclassification rates estimated from calibration data and prevent correction of the raw data. Our results illustrate the importance of testing for and quantifying observer error in species identification in wildlife census and population trend studies particularly when target species may be easily confused with other nontarget species.     

Classification and Error Estimation for Discrete Data

Discrete classification is common in Genomic Signal Processing applications, in particular in classification of discretized gene expression data, and in discrete gene expression prediction and the inference of boolean genomic regulatory networks. Once a discrete classifier is obtained from sample data, its performance must be evaluated through its classification error. In practice, error estimation methods must then be employed to obtain reliable estimates of the classification error based on the available data. Both classifier design and error estimation are complicated, in the case of Genomics, by the prevalence of small-sample data sets in such applications. This paper presents a broad review of the methodology of classification and error estimation for discrete data, in the context of Genomics, focusing on the study of performance in small sample scenarios, as well as asymptotic behavior.Key Words: Genomics, classification, error estimation, discrete histogram rule, sampling distribution, resubstitution, leave-one-out, ensemble methods, coefficient of determination.     

Ancient DNA extraction from bones and teeth

This method is designed to maximize recovery of PCR-amplifiable DNA from ancient bone and teeth specimens and at the same time to minimize co-extraction of substances that inhibit PCR. This is achieved by a combination of DNA extraction from bone powder using a buffer consisting solely of EDTA and proteinase K, and purification of the DNA by binding to silica in the presence of high concentrations of guanidinium thiocyanate. All steps are performed at room temperature (20-23 degrees C), thereby reducing further degradation of the already damaged and fragile ancient DNA and providing an optimal trade-off between DNA release and degradation. Furthermore, the purification step removes most of the various types of PCR inhibitors present in ancient bone samples, thereby optimizing the amount of ancient DNA available for subsequent enzymatic manipulation, such as PCR amplification. The protocol presented here allows DNA extraction from ancient bone and teeth with a minimum of working steps and equipment and yields DNA extracts within 2 working days.     

Serum Triglyceride: Method of Estimation and Levels in Normal Humans

Previously described glycerol methods for estimation of serum triglyceride were modified. Conditions during saponification and colour development were altered so as to minimize the possibility of glycerol loss. Using this modification, serum triglyceride was determined in 100 healthy men and women, aged 15 to 79 years. There was a log normal distribution. In both sexes the level increased up to the sixth decade and then decreased. In each decade men had higher levels than women. The geometric mean (and 95% limits) for men was 129 mg. % (68-248); for women, 105 mg. % (54-206); and for the entire group, 117 mg. % (59-233). Comparison of results from several laboratories using different methods showed wide variation in serum triglyceride levels.