Controlling for contamination in re-sequencing studies with a reproducible web-based phylogenetic approach

Polymorphism discovery is a routine application of next-generation sequencing technology where multiple samples are sent to a service provider for library preparation, subsequent sequencing, and bioinformatic analyses. The decreasing cost and advances in multiplexing approaches have made it possible to analyze hundreds of samples at a reasonable cost. However, because of the manual steps involved in the initial processing of samples and handling of sequencing equipment, cross-contamination remains a significant challenge. It is especially problematic in cases where polymorphism frequencies do not adhere to diploid expectation, for example, heterogeneous tumor samples, organellar genomes, as well as during bacterial and viral sequencing. In these instances, low levels of contamination may be readily mistaken for polymorphisms, leading to false results. Here we describe practical steps designed to reliably detect contamination and uncover its origin, and also provide new, Galaxy-based, readily accessible computational tools and workflows for quality control. All results described in this report can be reproduced interactively on the web as described at http://usegalaxy.org/contamination

Maternal age effect and severe germ-line bottleneck in the inheritance of human mitochondrial DNA

The manifestation of mitochondrial DNA (mtDNA) diseases depends on the frequency of heteroplasmy (the presence of several alleles in an individual), yet its transmission across generations cannot be readily predicted owing to a lack of data on the size of the mtDNA bottleneck during oogenesis. For deleterious heteroplasmies, a severe bottleneck may abruptly transform a benign (low) frequency in a mother into a disease-causing (high) frequency in her child. Here we present a high-resolution study of heteroplasmy transmission conducted on blood and buccal mtDNA of 39 healthy mother–child pairs of European ancestry (a total of 156 samples, each sequenced at ∼20,000× per site). On average, each individual carried one heteroplasmy, and one in eight individuals carried a disease-associated heteroplasmy, with minor allele frequency ≥1%. We observed frequent drastic heteroplasmy frequency shifts between generations and estimated the effective size of the germ-line mtDNA bottleneck at only ∼30–35 (interquartile range from 9 to 141). Accounting for heteroplasmies, we estimated the mtDNA germ-line mutation rate at 1.3 × 10−8 (interquartile range from 4.2 × 10−9 to 4.1 × 10−8) mutations per site per year, an order of magnitude higher than for nuclear DNA. Notably, we found a positive association between the number of heteroplasmies in a child and maternal age at fertilization, likely attributable to oocyte aging. This study also took advantage of droplet digital PCR (ddPCR) to validate heteroplasmies and confirm a de novo mutation. Our results can be used to predict the transmission of disease-causing mtDNA variants and illuminate evolutionary dynamics of the mitochondrial genome

Streamlined computational pipeline for genetic background characterization of genetically engineered mice based on next generation sequencing data

Abstract Background Genetically engineered mice (GEM) are essential tools for understanding gene function and disease modeling. Historically, gene targeting was first done in embryonic stem cells (ESCs) derived from the 129 family of inbred strains, leading to a mixed background or congenic mice when crossed with C57BL/6 mice. Depending on the number of backcrosses and breeding strategies, genomic segments from 129-derived ESCs can be introgressed into the C57BL/6 genome, establishing a unique genetic makeup that needs characterization in order to obtain valid conclusions from experiments using GEM lines. Currently, SNP genotyping is used to detect the extent of 129-derived ESC genome introgression into C57BL/6 recipients; however, it fails to detect novel/rare variants. Results Here, we present a computational pipeline implemented in the Galaxy platform and in BASH/R script to determine genetic introgression of GEM using next generation sequencing data (NGS), such as whole genome sequencing (WGS), whole exome sequencing (WES) and RNA-Seq. The pipeline includes strategies to uncover variants linked to a targeted locus, genome-wide variant visualization, and the identification of potential modifier genes. Although these methods apply to congenic mice, they can also be used to describe variants fixed by genetic drift. As a proof of principle, we analyzed publicly available RNA-Seq data from five congenic knockout (KO) lines and our own RNA-Seq data from the Sall2 KO line. Additionally, we performed target validation using several genetics approaches. Conclusions We revealed the impact of the 129-derived ESC genome introgression on gene expression, predicted potential modifier genes, and identified potential phenotypic interference in KO lines. Our results demonstrate that our new approach is an effective method to determine genetic introgression of GEM

A mouse systems genetics approach reveals common and uncommon genetic modifiers of hepatic lysosomal enzyme activities and glycosphingolipids

Identification of genetic modulators of lysosomal enzyme activities and glycosphingolipids (GSLs) may facilitate the development of therapeutics for diseases in which they participate, in-cluding Lysosomal Storage Disorders (LSDs). To this end, we used a systems genetics approach: we measured 11 hepatic lysosomal enzymes and many of their natural substrates (GSLs), fol-lowed by modifier gene mapping by GWAS and transcriptomics associations in a panel of inbred strains. Unexpectedly, most GSLs showed no association between their levels and the enzyme ac-tivity that catabolizes them. Genomic mapping identified 30 shared predicted modifier genes between the enzymes and GSLs, which are clustered in three pathways and are associated with other diseases. Surprisingly, they are regulated by ten common transcription factors, and their majority by miRNA-340p. In conclusion, we have identified novel regulators of GSL metabolism, which may serve as therapeutic targets for LSDs and may suggest the involvement of GSL metab-olism in other pathologies

Encuentro de Ciencias Básicas 2: retos frente a la deserción

Pensar en la deserción –en adelante abandono– universitaria conlleva contemplar un espectro de factores que inciden en la misma, de los cuales se destacan condiciones académicas previas, el factor económico –concluyente en la continuidad o no de los estudios superiores–; aspectos sociales, culturales y familiares que influyen en la escogencia del programa; el sentido de pertenencia frente a la institución en la cual se va a formar el estudiante, y otros tantos, que permiten afirmar que la deserción en educación superior no se debe solo a la pérdida de las asignaturas pertenecientes a las ciencias básicas. Es por esto que en su segunda versión el Encuentro de Ciencias Básicas centró sus esfuerzos en evidenciar los hechos que inciden en la transición colegio-universidad, como un elemento determinante en el abandono. Este tópico fue resultado del primer encuentro, donde los profesores-investigadores cuestionaron los esfuerzos que hacen las universidades, los cuales se enfocan en solucionar problemáticas asociadas con la formación secundaria. Por tanto, se hizo imperante indagar sobre qué se está haciendo para estudiar, analizar y estructurar propuestas metodológicas que faciliten un tránsito menos complejo para los estudiantes que dejaron su colegio aún siendo niños y los recibe la universidad como adultos. Así pues, estas memorias se convierten en un insumo de gran valor para las universidades y los colegios que se estén cuestionado sobre sus procesos de formación, donde no solo se preocupen por el carácter propedéutico de la misma, sino también se repiense la importancia de la preparación para la vida universitaria, o qué metodologías son las más asertivas para la recepción de los estudiantes de los primeros semestres universitarios, que son, evidentemente, la población más vulnerable al fenómeno de abandono de la educación superior.Acompañamiento, una propuesta de evaluación formativa. Un ejercicio de modelación matemática, aplicando los conceptos del álgebra lineal con estudiantes de la Facultad de Ingeniería y el programa de Economía de la Universidad Católica de Colombia En busca de la relación entre ciencias básicas y deserción en la educación superior colombiana Experiencia de transición aritmética-álgebra con estudiantes de primer semestre de la Universidad Católica de Colombia Experiencias de aprendizaje en proyectos de aula: una estrategia para la retención universitaria MOEBIUS: hacia una didáctica de la transición enseñanza media superior-universidad Formulación de una estrategia para la enseñanza del concepto de la derivada a partir de los conocimientos previos y su impacto en la disminución de la deserción escolar La indagación en proyectos transversales como estrategia pedagógica en la mediación entre la educación media y la universitaria. La tutoría: factor que motiva el rendimiento académico en estudiantes en la transición colegio-universidad. Curso de acogida para estudiantes neotomasinos de la División de Ingenierías, fundamentado en el razonamiento, el pensamiento lógico-matemático y el trabajo colaborativo Propuesta interinstitucional de acompañamiento académico a partir del diseño e implementación de pruebas diagnósticas. Representaciones del movimiento parabólico para niveles diferenciados de educación media a superior