Base-calling of automated sequencer traces using phred. I. Accuracy assessment.

By B. Ewing, LaDeana W. Hillier, Michael C. Wendl, Philip Green. 1998

The availability of massive amounts of DNA sequence information has begun to revolutionize the practice of biology. As a result, current large-scale sequencing output, while impressive, is not adequate to keep pace with growing demand and, in particular, is far short of what will be required to obtain the 3-billion-base human genome sequence by the target date of 2005. To reach this goal, improved automation will be essential, and it is particularly important that human involvement in sequence data processing be significantly reduced or eliminated. Progress in this respect will require both improved accuracy of the data processing software and reliable accuracy measures to reduce the need for human involvement in error correction and make human review more efficient. Here, we describe one step toward that goal: a base-calling program for automated sequencer traces, phred, with improved accuracy. phred appears to be the first base-calling program to achieve a lower error rate than the ABI software, averaging 40%-50% fewer errors in the data sets examined independent of position in read, machine running conditions, or sequencing chemistry.

How researchers see this paper

The researchers whose trusted networks rank it highest, strongest advocate first. Every researcher gets a different order.

Citations in the corpus

CADD: predicting the deleteriousness of variants throughout the human genome (2018, background): “log10-derived, (6)) rank score based on the genome-wide distribution of scores for all ∼9 billion potential SNVs, the set of all three non-reference alleles at each position of the reference assembly.”
Canu: scalable and accurate long-read assembly via adaptive k-mer weighting and repeat separation (2016, background): “An ideal assembly has high continuity, low breakpoints, and high base accuracy, with 99.99% (Phred QV40) (Ewing and Green 1998) commonly defined as the minimum quality for a “ finished ” sequence (Felsenfeld et al. 1999; Schmutz et al. 2004).”
Testing the effect of palaeodrainages versus habitat stability on genetic divergence in riverine systems: study of a Neotropical fish of the Brazilian coastal Atlantic Forest (2015, methodology): “Two mitochondrial genes, cytochrome oxidase I (COI) and NADH dehydrogenase 2 (ND2), were amplified and sequenced following standard protocols (for details see Thomaz et al., 2010), with forward and reverse chromatogram assembled and visualized using Phred/Phrap/Consed (Ewing et al., 1998; Gordon et ”
Biodegradation of textile azo dyes by a facultative Staphylococcus arlettae strain VN-11 using a sequential microaerophilic/aerobic process. (2009, methodology): “Partial 16S rRNA sequences obtained from isolates were assembled in a contig using the phred/Phrap/CONSED program (Godon et al., 1997; Ewing et al., 1998).”
SPAdes: A New Genome Assembly Algorithm and Its Applications to Single-Cell Sequencing (2012, background): “(4) Stage 4 (contig construction) was well studied in the context of Sanger sequencing (Ewing et al., 1998).”
Protein interaction mapping: a Drosophila case study. (2005, methodology): “5 and 3 sequences were then filtered by using PHRED (Ewing and Green 1998; Ewing et al. 1998) and masking ALU repeats.”
Minimus: a fast, lightweight genome assembler (2007, methodology): “The first generation of assemblers, including TIGR Assembler [1], phrap [2], and CAP3 [3], were able to assemble smallto medium-sized bacterial genomes, often requiring several weeks of computer time on the fastest computers then available.”
EzTaxon: a web-based tool for the identification of prokaryotes based on 16S ribosomal RNA gene sequences. (2007, methodology): “The PHRED program was used for the base-calling and trimming of uploaded tracer files (Ewing & Green, 1998; Ewing et al., 1998).”
Hd1, a Major Photoperiod Sensitivity Quantitative Trait Locus in Rice, Is Closely Related to the Arabidopsis Flowering Time Gene CONSTANS (2000, methodology): “The raw sequence data of the PAC insert were scored with phred software (Ewing et al., 1998) and then assembled with phrap/cross-match software to make a contiguous nucleotide sequence (Ewing and Green, 1998).”