Multigene amplification and massively parallel sequencing for cancer mutation discovery

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.

Stenberg, Johan (författare)
Fredriksson, Simon (författare)
Welch, Katrina (författare)
Zhang, Michael (författare)
Nilsson, MatsUppsala universitet,Institutionen för genetik och patologi(Swepub:uu)matsnils (författare)
Bicknell, David (författare)
Bodmer, Walter F. (författare)
Davis, Ronald W. (författare)
Ji, Hanlee (författare)
Uppsala universitetInstitutionen för genetik och patologi (creator_code:org_t)

Ingår i:Proceedings of the National Academy of Sciences of the United States of America: Proceedings of the National Academy of Sciences104:22, s. 9387-93920027-84241091-6490

Av författaren/redakt...: Dahl, Fredrik; Stenberg, Johan; Fredriksson, Sim ...; Welch, Katrina; Zhang, Michael; Nilsson, Mats; visa fler...; Bicknell, David; Bodmer, Walter F ...; Davis, Ronald W.; Ji, Hanlee; visa färre...

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