Measuring sequencer size bias using REcount: a novel method for highly accurate Illumina sequencing-based quantification.
Genome Biol
; 20(1): 85, 2019 04 29.
Article
em En
| MEDLINE
| ID: mdl-31036053
ABSTRACT
Quantification of DNA sequence tags from engineered constructs such as plasmids, transposons, or other transgenes underlies many functional genomics measurements. Typically, such measurements rely on PCR followed by next-generation sequencing. However, PCR amplification can introduce significant quantitative error. We describe REcount, a novel PCR-free direct counting method. Comparing measurements of defined plasmid pools to droplet digital PCR data demonstrates that REcount is highly accurate and reproducible. We use REcount to provide new insights into clustering biases due to molecule length across different Illumina sequencers and illustrate the impacts on interpretation of next-generation sequencing data and the economics of data generation.
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Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Enzimas de Restrição do DNA
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Sitios de Sequências Rotuladas
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Técnicas Genéticas
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article