MixDeR: A SNP mixture deconvolution workflow for forensic genetic genealogy.

The generation of forensic DNA profiles consisting of single nucleotide polymorphisms (SNPs) is now being facilitated by wider adoption of next-generation sequencing (NGS) methods in casework laboratories. At the same time, and in part because of this advance, there is an intense focus on the generation of SNP profiles from evidentiary specimens for so-called forensic or investigative genetic genealogy (FGG or IGG) applications. However, FGG methods are constrained by the algorithms for genealogical database searches, which were designed for use with single-source profiles, and the fact that many forensic samples are mixtures.

Sequencing-induced artefacts in NGS STR data.

Significant progress has been made in recent years in the development of techniques for Next Generation Sequencing (NGS), or Massively Parallel Sequencing (MPS), of forensically relevant short tandem repeat (STR) loci. However, as these technologies are investigated and adopted by forensic laboratories, new challenges unfold that require further scrutiny. In the analysis of DNA profiles generated using the MiSeq FGx sequencing system, we have observed noise sequences with relatively high readcounts that are challenging to distinguish from genuine alleles.

MPSproto: An extension of EuroForMix to evaluate MPS-STR mixtures.

We have developed MPSproto as an extension of EuroForMix to improve handling of stutter artefacts and other typing errors that commonly occur in MPS-STR data. MPSproto implements two models for read depth: gamma and negative binomial. It differs from EuroForMix in that calibration is required before mixtures are interpreted.

The long-acting amylin/calcitonin receptor agonist ZP5461 suppresses food intake and body weight in male rats.

The peptide hormone amylin reduces food intake and body weight and is an attractive candidate target for novel pharmacotherapies to treat obesity. However, the short half-life of native amylin and amylin analogs like pramlintide limits these compounds' potential utility in promoting sustained negative energy balance. Here, we evaluate the ability of the novel long-acting amylin/calcitonin receptor agonist ZP5461 to reduce feeding and body weight in rats, and also test the role of calcitonin receptors (CTRs) in the dorsal vagal complex (DVC) of the hindbrain in the energy balance effects of chronic ZP5461 administration.

Modeling allelic analyte signals for aSTRs in NGS DNA profiles.

We describe an adaption of Bright et al.'s work modeling peak height variability in CE-DNA profiles to the modeling of allelic aSTR (autosomal short tandem repeats) read counts from NGS-DNA profiles, specifically for profiles generated from the ForenSeq™ DNA Signature Prep Kit, DNA Primer Mix B. Bright et al.

Variability and additivity of read counts for aSTRs in NGS DNA profiles.

There has been an increase in the number of laboratories and researchers adopting new sequencing technologies, known as next-generation sequencing (NGS). An understanding of the behaviour of NGS DNA profiles is needed to enable for the development of probabilistic genotyping methods for the interpretation of such profiles. In this work, we investigate NGS analyte signal variation, specifically heterozygous balance and stutter variability from profiles generated using the ForenSeq™ DNA Signature Prep Kit, DNA Primer Mix B.

An examination of STR nomenclatures, filters and models for MPS mixture interpretation.

The increased interest in the use of Massively Parallel Sequencing (MPS) technologies to type traditional autosomal STR markers raises multiple questions regarding interpretation of the results via probabilistic genotyping. To begin to address some of those questions, we examined the effects of using differing degrees of sequence information, pre-filtering, and data modeling to interpret complex MPS-STR mixtures in a probabilistic genotyping software. Sixty ForenSeq typing results for mixtures of from two to four contributors were: 1) represented using three separate formats that captured different degrees of sequence information, and 2) were analyzed using three different filtering approaches prior to probabilistic interpretation.

Validation of NGS for mitochondrial DNA casework at the FBI Laboratory.

As a first step towards integrating next generation sequencing (NGS) technology into the FBI Laboratory's operational casework, the PowerSeq™ CRM Nested System, an NGS-based mitochondrial DNA (mtDNA) control region assay, was developmentally and internally validated. The validation studies were conducted in accordance with the Scientific Working Group on DNA Analysis Methods (SWGDAM) Validation Guidelines for Forensic DNA Analysis Methods, and the FBI's Quality Assurance Standards (QAS) for Forensic DNA Testing Laboratories. The assay was shown to be highly reproducible, with variant frequencies across intra and inter-run replicates of the same sample differing, on average, by just 0.

Modeling household energy consumption and adoption of energy efficient technology.

This study develops a model to study household energy use behavior that can impose common preferences for feasible demand estimation with multiple discrete technology choices and multiple continuous energy consumption uses. The model imposes fixed proportions production and additivity of uses for plausible estimation feasibility while adopting a second-order translog flexible functional form to focus on flexibility in identification of consumer preferences that determine interactions among energy uses and between short-run and long-run choices. Using a unique household-level dataset from California, the model is applied to estimate short-run household demand for electricity and natural gas and the long-run technology choices with respect to clothes washing, water heating, space heating, and clothes drying.

Trans-Graft Protection Against Pierce's Disease Mediated by Transgenic Grapevine Rootstocks.

A field study showed that transgenic grapevine rootstocks can provide trans-graft-mediated protection to a wild type scion against Pierce's disease (PD) development. We individually field-tested two distinct strategies. The first expressed a chimeric antimicrobial protein (CAP) that targeted the functionality of the lipopolysaccharide (LPS) surface of (), the causative agent of PD.

A closer look at Verogen's Forenseq™ DNA Signature Prep kit autosomal and Y-STR data for streamlined analysis of routine reference samples.

Massively parallel sequencing (MPS) provides forensic DNA laboratories an option to overcome the limitations associated with CE and current STR assays. Verogen's MPS Forenseq DNA Signature kit concomitantly amplifies 27 autosomal, 7 X-, and 24 Y-STRs. In addition, 94 identity, 56 ancestry, and 22 phenotypic-informative SNPs are included for a total of over 200 markers in one multiplex.

Use of the LUS in sequence allele designations to facilitate probabilistic genotyping of NGS-based STR typing results.

Some of the expected advantages of next generation sequencing (NGS) for short tandem repeat (STR) typing include enhanced mixture detection and genotype resolution via sequence variation among non-homologous alleles of the same length. However, at the same time that NGS methods for forensic DNA typing have advanced in recent years, many caseworking laboratories have implemented or are transitioning to probabilistic genotyping to assist the interpretation of complex autosomal STR typing results. Current probabilistic software programs are designed for length-based data, and were not intended to accommodate sequence strings as the product input.

Characterization of signalling and regulation of common calcitonin receptor splice variants and polymorphisms.

The calcitonin receptor (CTR) is a class B G protein-coupled receptor that is a therapeutic target for the treatment of hypercalcaemia of malignancy, Paget's disease and osteoporosis. In primates, the CTR is subject to alternative splicing, with a unique, primate-specific splice variant being preferentially expressed in reproductive organs, lung and kidney. In addition, humans possess a common non-synonymous single-nucleotide polymorphism (SNP) encoding a proline/leucine substitution in the C-terminal tail.

Internal validation of STRmix™ for the interpretation of single source and mixed DNA profiles.

The interpretation of DNA evidence can entail analysis of challenging STR typing results. Genotypes inferred from low quality or quantity specimens, or mixed DNA samples originating from multiple contributors, can result in weak or inconclusive match probabilities when a binary interpretation method and necessary thresholds (such as a stochastic threshold) are employed. Probabilistic genotyping approaches, such as fully continuous methods that incorporate empirically determined biological parameter models, enable usage of more of the profile information and reduce subjectivity in interpretation.

Performance and concordance of the ForenSeq™ system for autosomal and Y chromosome short tandem repeat sequencing of reference-type specimens.

Though the utility of next-generation sequencing (NGS) technologies for forensic short tandem repeat (STR) typing has been evident for several years, commercially available assays and software solutions developed specifically to meet forensic needs have only recently become available. One of these, the ForenSeq™ DNA Signature Prep Kit (Illumina, Inc.) sequences 27 autosomal STR (aSTR) and 24 Y chromosome STR (Y-STR) loci (concurrent with additional nuclear markers) per multiplexed sample, with automated secondary and tertiary analyses of the data accomplished via the associated ForenSeq™ Universal Analysis Software (UAS).

First autochtonous infection of a dog with Oslerus (Filaroides) osleri in the Czech Republic.

This case report describes an infection with O. osleri in a 10-month-old intact female Miniature German Spitz that presented with a 3-month history of progressive cough. Diagnosis was based upon visualization of characteristic lesions during bronchoscopy.

The mitochondrial landscape of African Americans: An examination of more than 2500 control region haplotypes from 22 U.S. locations.

The mitochondrial DNA (mtDNA) control region (16024-576) was Sanger-sequenced for a total of 2563 self-identified African Americans, using automated processing techniques and data review standards exceeding guidelines for forensic applications. Genetic diversity ranged from 0.9952 to 0.

Mitochondrial DNA heteroplasmy in the emerging field of massively parallel sequencing.

Long an important and useful tool in forensic genetic investigations, mitochondrial DNA (mtDNA) typing continues to mature. Research in the last few years has demonstrated both that data from the entire molecule will have practical benefits in forensic DNA casework, and that massively parallel sequencing (MPS) methods will make full mitochondrial genome (mtGenome) sequencing of forensic specimens feasible and cost-effective. A spate of recent studies has employed these new technologies to assess intraindividual mtDNA variation.

Effects of Roux-en-Y gastric bypass on fasting and postprandial inflammation-related parameters in obese subjects with normal glucose tolerance and in obese subjects with type 2 diabetes.

Background: Obesity is characterized by low grade inflammation and an altered secretion of inflammatory cytokines from the adipose tissue. Weight loss has shown to reduce inflammation; however, changes in cytokine profiles during massive weight loss are not well described. The present study explored the hypothesis that Roux-en-Y gastric bypass (RYGB) reduces circulating levels of pro-inflammatory cytokines, while increasing anti-inflammatory cytokines in obese subjects with type 2 diabetes (T2D) and in obese normal glucose tolerant (NGT) subjects.

Full mtGenome reference data: development and characterization of 588 forensic-quality haplotypes representing three U.S. populations.

Though investigations into the use of massively parallel sequencing technologies for the generation of complete mitochondrial genome (mtGenome) profiles from difficult forensic specimens are well underway in multiple laboratories, the high quality population reference data necessary to support full mtGenome typing in the forensic context are lacking. To address this deficiency, we have developed 588 complete mtGenome haplotypes, spanning three U.S.

Short tandem repeat typing on the 454 platform: strategies and considerations for targeted sequencing of common forensic markers.

To investigate the feasibility of next generation sequencing technology (NGS) for the multiplex detection and sequence production of short tandem repeats (STRs) from degraded and low DNA quantity samples, standard polymerase chain reaction amplification methods were used to enrich for commonly employed STR markers. Samples were amplified with two multiplexing strategies: a multiplex containing thirteen miniSTR markers and a series of multiplexes containing four miniSTR markers each. Each sample multiplex was barcoded with a sample-specific multiplex identifier for subsequent parallel tagged sequencing on the GS Junior System (454 Life Sciences, a Roche company, Branford, CT).

Development of forensic-quality full mtGenome haplotypes: success rates with low template specimens.

Forensic mitochondrial DNA (mtDNA) testing requires appropriate, high quality reference population data for estimating the rarity of questioned haplotypes and, in turn, the strength of the mtDNA evidence. Available reference databases (SWGDAM, EMPOP) currently include information from the mtDNA control region; however, novel methods that quickly and easily recover mtDNA coding region data are becoming increasingly available. Though these assays promise to both facilitate the acquisition of mitochondrial genome (mtGenome) data and maximize the general utility of mtDNA testing in forensics, the appropriate reference data and database tools required for their routine application in forensic casework are lacking.

A high-throughput Sanger strategy for human mitochondrial genome sequencing.

Background: A population reference database of complete human mitochondrial genome (mtGenome) sequences is needed to enable the use of mitochondrial DNA (mtDNA) coding region data in forensic casework applications. However, the development of entire mtGenome haplotypes to forensic data quality standards is difficult and laborious. A Sanger-based amplification and sequencing strategy that is designed for automated processing, yet routinely produces high quality sequences, is needed to facilitate high-volume production of these mtGenome data sets.