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Related Articles Profiling model T cell metagenomes with short reads.
Bioinformatics. 2009 Jan 9;
Authors: Warren RL, Nelson BH, Holt RA
MOTIVATION: T cell receptor (TCR) diversity in peripheral blood has not yet been fully profiled with sequence level resolution. Each T cell clonotype expresses a unique receptor, generated by somatic recombination of TCR genes and the enormous potential for T cell diversity makes repertoire analysis challenging. We developed a sequencing approach and assembly software (immuno-SSAKE or iSSAKE) for profiling T cell metagenomes using short reads from the massively parallel sequencing platforms. RESULTS: Models of sequence diversity for the TCR beta-chain CDR3 region were built using empirical data and used to simulate, at random, distinct TCR clonotypes at 1-20 parts per million. Using simulated TCRbeta (sTCRbeta) sequences, we randomly created 20 million 36nt reads having 1%-2% random error, 20 million 42nt or 50nt reads having 1% random error and 20 million 36nt reads with 1% error modeled on real short read data. Reads aligning to the end of known TCR variable (V) genes and having consecutive unmatched bases in the adjacent CDR3 were used to seed iSSAKE de novo assemblies of CDR3. With assembled 36nt reads we detect over 51% and 63% of rare (1 ppm) clonotypes using a random or modeled error distribution, respectively. We detect over 99% of more abundant clonotypes (6 ppm or higher) using either error distribution. Longer reads improve sensitivity, with assembled 42nt and 50nt reads identifying 82.0% and 94.7% of rare 1 ppm clonotypes, respectively. Our approach illustrates the feasibility of complete profiling of the TCR repertoire using new massively parallel short read sequencing technology. AVAILABILITY: ftp://ftp.bcgsc.ca/supplementary/iSSAKE. CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary methods and data available at Bioinformatics online and at ftp://ftp.bcgsc.ca/supplementary/iSSAKE.
PMID: 19136549 [PubMed - as supplied by publisher]
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Related Articles Profiling model T cell metagenomes with short reads.
Bioinformatics. 2009 Jan 9;
Authors: Warren RL, Nelson BH, Holt RA
MOTIVATION: T cell receptor (TCR) diversity in peripheral blood has not yet been fully profiled with sequence level resolution. Each T cell clonotype expresses a unique receptor, generated by somatic recombination of TCR genes and the enormous potential for T cell diversity makes repertoire analysis challenging. We developed a sequencing approach and assembly software (immuno-SSAKE or iSSAKE) for profiling T cell metagenomes using short reads from the massively parallel sequencing platforms. RESULTS: Models of sequence diversity for the TCR beta-chain CDR3 region were built using empirical data and used to simulate, at random, distinct TCR clonotypes at 1-20 parts per million. Using simulated TCRbeta (sTCRbeta) sequences, we randomly created 20 million 36nt reads having 1%-2% random error, 20 million 42nt or 50nt reads having 1% random error and 20 million 36nt reads with 1% error modeled on real short read data. Reads aligning to the end of known TCR variable (V) genes and having consecutive unmatched bases in the adjacent CDR3 were used to seed iSSAKE de novo assemblies of CDR3. With assembled 36nt reads we detect over 51% and 63% of rare (1 ppm) clonotypes using a random or modeled error distribution, respectively. We detect over 99% of more abundant clonotypes (6 ppm or higher) using either error distribution. Longer reads improve sensitivity, with assembled 42nt and 50nt reads identifying 82.0% and 94.7% of rare 1 ppm clonotypes, respectively. Our approach illustrates the feasibility of complete profiling of the TCR repertoire using new massively parallel short read sequencing technology. AVAILABILITY: ftp://ftp.bcgsc.ca/supplementary/iSSAKE. CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary methods and data available at Bioinformatics online and at ftp://ftp.bcgsc.ca/supplementary/iSSAKE.
PMID: 19136549 [PubMed - as supplied by publisher]
More...