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We have developed a procedure for massively parallel resequencing of multiple human genes by combining a highly multiplexed and target-specific amplification process with a high-throughput parallel sequencing technology. The amplification process is based on oligonucleotide constructs, called selectors, that guide the circularization of specific DNA target regions. Subsequently, the circularized target sequences are amplified in multiplex and analyzed by using a highly parallel sequencing-by-synthesis technology. As a proof-of-concept study, we demonstrate parallel resequencing of 10 cancer genes covering 177 exons with average sequence coverage per sample of 93%. Seven cancer cell lines and one normal genomic DNA sample were studied with multiple mutations and polymorphisms identified among the 10 genes. Mutations and polymorphisms in the TP53 gene were confirmed by traditional sequencing.

Original publication

DOI

10.1073/pnas.0702165104

Type

Journal article

Journal

Proc Natl Acad Sci U S A

Publication Date

29/05/2007

Volume

104

Pages

9387 - 9392

Keywords

Base Sequence, Cell Line, Tumor, Genes, Neoplasm, Genetic Testing, Humans, Molecular Sequence Data, Mutation, Neoplasms, Nucleic Acid Amplification Techniques, Tumor Suppressor Protein p53