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Replicate exome-sequencing in a multiple-generation family: improved interpretation of next-generation sequencing data

Overview of attention for article published in BMC Genomics, November 2015
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Title
Replicate exome-sequencing in a multiple-generation family: improved interpretation of next-generation sequencing data
Published in
BMC Genomics, November 2015
DOI 10.1186/s12864-015-2107-y
Pubmed ID
Authors

Praveen F. Cherukuri, Valerie Maduro, Karin V. Fuentes-Fajardo, Kevin Lam, David R. Adams, Cynthia J. Tifft, James C. Mullikin, William A. Gahl, Cornelius F. Boerkoel

Abstract

Whole-exome sequencing (WES) is rapidly evolving into a tool of choice for rapid, and inexpensive identification of molecular genetic lesions within targeted regions of the human genome. While biases in WES coverage of nucleotides in targeted regions are recognized, it is not well understood how repetition of WES improves the interpretation of sequencing results in a clinical diagnostic setting. To address this, we compared independently generated exome-capture of six individuals from three-generations sequenced in triplicate. This generated between 48x-86x mean target depth of high-quality mapped bases (>Q20) for each technical replicate library. Cumulatively, we achieved 179 - 208x average target coverage for each individual in the pedigree. Using this experimental design, we evaluated stochastics in WES interpretation, genotyping sensitivity, and accuracy to detect de novo variants. In this study, we show that repetition of WES improved the interpretation of the capture target regions after aggregating the data (93.5 - 93.9 %). Compared to 81.2 - 89.6 % (50.2-55.4 Mb of 61.7 M) coverage of targeted bases at ≥20x in the individual technical replicates, the aggregated data covered 93.5 - 93.9 % of targeted bases (57.7 - 58.0 of 61.7 M) at ≥20x threshold, suggesting a 4.3 - 12.7 % improvement in coverage. Each individual's aggregate dataset recovered 3.4 - 6.4 million bases within variable targeted regions. We uncovered technical variability (2-5 %) inherent to WES technique. We also show improved interpretation in assessing clinically important regions that lack interpretation under current conditions, affecting 12-16 of the 56 genes recommended for secondary analysis by American College of Medical Genetics (ACMG). We demonstrate that comparing technical replicate WES datasets and their derived aggregate data can effectively address overall WES genotyping discrepancies. We describe a method to evaluate the reproducibility and stochastics in exome library preparation, and delineate the advantages of aggregating the data derived from technical replicates. The implications of this study are directly applicable to improved experimental design and provide an opportunity to rapidly, efficiently, and accurately arrive at reliable candidate nucleotide variants.

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Mendeley readers

Mendeley readers

The data shown below were compiled from readership statistics for 23 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
United States 1 4%
Unknown 22 96%

Demographic breakdown

Readers by professional status Count As %
Researcher 9 39%
Student > Ph. D. Student 3 13%
Other 2 9%
Professor 1 4%
Student > Master 1 4%
Other 1 4%
Unknown 6 26%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 11 48%
Agricultural and Biological Sciences 4 17%
Neuroscience 1 4%
Medicine and Dentistry 1 4%
Unknown 6 26%
Attention Score in Context

Attention Score in Context

This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 01 December 2015.
All research outputs
#14,241,439
of 22,833,393 outputs
Outputs from BMC Genomics
#5,703
of 10,655 outputs
Outputs of similar age
#202,236
of 386,751 outputs
Outputs of similar age from BMC Genomics
#233
of 388 outputs
Altmetric has tracked 22,833,393 research outputs across all sources so far. This one is in the 35th percentile – i.e., 35% of other outputs scored the same or lower than it.
So far Altmetric has tracked 10,655 research outputs from this source. They receive a mean Attention Score of 4.7. This one is in the 42nd percentile – i.e., 42% of its peers scored the same or lower than it.
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We're also able to compare this research output to 388 others from the same source and published within six weeks on either side of this one. This one is in the 33rd percentile – i.e., 33% of its contemporaries scored the same or lower than it.