276 research outputs found
Differential meta-analysis of RNA-seq data from multiple studies
High-throughput sequencing is now regularly used for studies of the
transcriptome (RNA-seq), particularly for comparisons among experimental
conditions. For the time being, a limited number of biological replicates are
typically considered in such experiments, leading to low detection power for
differential expression. As their cost continues to decrease, it is likely that
additional follow-up studies will be conducted to re-address the same
biological question. We demonstrate how p-value combination techniques
previously used for microarray meta-analyses can be used for the differential
analysis of RNA-seq data from multiple related studies. These techniques are
compared to a negative binomial generalized linear model (GLM) including a
fixed study effect on simulated data and real data on human melanoma cell
lines. The GLM with fixed study effect performed well for low inter-study
variation and small numbers of studies, but was outperformed by the
meta-analysis methods for moderate to large inter-study variability and larger
numbers of studies. To conclude, the p-value combination techniques illustrated
here are a valuable tool to perform differential meta-analyses of RNA-seq data
by appropriately accounting for biological and technical variability within
studies as well as additional study-specific effects. An R package metaRNASeq
is available on the R Forge
Transcriptomic evaluation of the nicotinic acetylcholine receptor pathway in levamisole-resistant and -sensitive Oesophagostomum dentatum
Nematode anthelminthic resistance is widespread for the 3 major drug classes commonly used in agriculture: benzamidazoles, macrocyclic lactones, and nicotinic agonists e.g. levamisole. In parasitic nematodes the genetics of resistance is unknown other than to the benzimidazoles which primarily involve a single gene. In previous work with a levamisole resistant Oesophagostomum dentatum isolate, the nicotinic acetylcholine receptor (nAChR) exhibited decreased levamisole sensitivity. Here, using a transcriptomic approach on the same isolate, we investigate whether that decreased nAChR sensitivity is achieved via a 1-gene mechanism involving 1 of 27 nAChR pathway genes. 3 nAChR receptor subunit genes exhibited ≥2-fold change in transcript abundance: acr-21 and acr-25 increased, and unc-63 decreased. 4 SNPs having a ≥2-fold change in frequency were also identified. These data suggest that resistance is likely polygenic, involving modulated abundance of multiple subunits comprising the heteropentameric nAChR, and is not due to a simple 1-gene mechanism
Essential guidelines for computational method benchmarking
In computational biology and other sciences, researchers are frequently faced
with a choice between several computational methods for performing data
analyses. Benchmarking studies aim to rigorously compare the performance of
different methods using well-characterized benchmark datasets, to determine the
strengths of each method or to provide recommendations regarding suitable
choices of methods for an analysis. However, benchmarking studies must be
carefully designed and implemented to provide accurate, unbiased, and
informative results. Here, we summarize key practical guidelines and
recommendations for performing high-quality benchmarking analyses, based on our
experiences in computational biology.Comment: Minor update
Comparison of different differential expression analysis tools for rna-seq data
In molecular biology research, RNA-seq is a relatively new method for transcriptome profiling. It utilizes the next generation sequencing technology to provide huge amount information about the variety and abundance of RNA present in an organism of interest at a specific state and a given time. One of the most important tasks of RNA-seq analysis is finding genes that are expressed differently in different subject groups. A lot of differential expression analysis tools for RNA-seq have been developed, but there is no golden standard in this field. In this research, four commonly used tools (DESeq, edgeR, limma, and cuffdiff) are studied by comparing their performances in the normalization of different subject group data, and also in the sensitivity and specificity of selection of genes with differential expression. In addition, their performances on genes which only express in one condition are compared. The data used are SEQC and melanoma. The result shows that in differential expression analysis, DESeq is slightly better than other tools in normalization, while DESeq, edgeR, and limma, in general, display good sensitivity and specificity, and limma outputs less false positive predictions. In cases where genes of interest are absent in one of the conditions, limma has the best performance
Optimization of miRNA-seq data preprocessing.
The past two decades of microRNA (miRNA) research has solidified the role of these small non-coding RNAs as key regulators of many biological processes and promising biomarkers for disease. The concurrent development in high-throughput profiling technology has further advanced our understanding of the impact of their dysregulation on a global scale. Currently, next-generation sequencing is the platform of choice for the discovery and quantification of miRNAs. Despite this, there is no clear consensus on how the data should be preprocessed before conducting downstream analyses. Often overlooked, data preprocessing is an essential step in data analysis: the presence of unreliable features and noise can affect the conclusions drawn from downstream analyses. Using a spike-in dilution study, we evaluated the effects of several general-purpose aligners (BWA, Bowtie, Bowtie 2 and Novoalign), and normalization methods (counts-per-million, total count scaling, upper quartile scaling, Trimmed Mean of M, DESeq, linear regression, cyclic loess and quantile) with respect to the final miRNA count data distribution, variance, bias and accuracy of differential expression analysis. We make practical recommendations on the optimal preprocessing methods for the extraction and interpretation of miRNA count data from small RNA-sequencing experiments
Essential guidelines for computational method benchmarking
In computational biology and other sciences, researchers are frequently faced with a choice between several computational methods for performing data analyses. Benchmarking studies aim to rigorously compare the performance of different methods using well-characterized benchmark datasets, to determine the strengths of each method or to provide recommendations regarding suitable choices of methods for an analysis. However, benchmarking studies must be carefully designed and implemented to provide accurate, unbiased, and informative results. Here, we summarize key practical guidelines and recommendations for performing high-quality benchmarking analyses, based on our experiences in computational biology
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