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Allelotyping by massively parallel pyrosequencing of SNP‐carrying trinucleotide threads
Author(s) -
Pettersson Erik,
Zajac Pawel,
Ståhl Patrik L.,
Jacobsson Josefin A.,
Fredriksson Robert,
Marcus Claude,
Schiöth Helgi B.,
Lundeberg Joakim,
Ahmadian Afshin
Publication year - 2008
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.20655
Subject(s) - biology , pyrosequencing , massively parallel , snp , genetics , massive parallel sequencing , parallel computing , computational biology , gene , single nucleotide polymorphism , dna sequencing , computer science , genotype
Here we present an approach for allelotyping combining the multiplexing features of the trinucleotide threading (TnT) method with pooling of genomic DNA and massively parallel pyrosequencing, enabling reliable allele frequency estimation in large cohorts. The approach offers several benefits as compared to array‐based methods and allows undertaking highly complex studies without compromising accuracy, while keeping the workload to a minimum. This proof‐of‐concept study involves formation of trinucleotide threads, targeting a total of 147 single‐nucleotide polymorphisms (SNPs) related to obesity and cancer, for multiplex amplification and allele extraction from a pool of 462 genomes, followed by massively parallel pyrosequencing. Approximately 177k reads were approved, identified, and assigned to SNP‐carrying threads rendering representative allele frequencies in the cohort. Hum Mutat 29(2), 323–329, 2008. © 2007 Wiley‐Liss, Inc.