Validation of whole genome amplification for analysis of the p53 tumor suppressor gene in limited amounts of tumor samples

Hasmats, JohannaKTH,Genteknologi,Science for Life Laboratory, SciLifeLab,Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden (författare)

QC 20120928
AuthorCount:9;
Personalized cancer treatment requires molecular characterization of individual tumor biopsies. These samples are frequently only available in limited quantities hampering genomic analysis. Several whole genome amplification (WGA) protocols have been developed with reported varying representation of genomic regions post amplification. In this study we investigate region dropout using a 929 polymerase based WGA approach. DNA from 123 lung cancers specimens and corresponding normal tissue were used and evaluated by Sanger sequencing of the p53 exons 5-8. To enable comparative analysis of this scarce material, WGA samples were compared with unamplified material using a pooling strategy of the 123 samples. In addition, a more detailed analysis of exon 7 amplicons were performed followed by extensive cloning and Sanger sequencing. Interestingly, by comparing data from the pooled samples to the individually sequenced exon 7, we demonstrate that mutations are more easily recovered from WGA pools and this was also supported by simulations of different sequencing coverage. Overall this data indicate a limited random loss of genomic regions supporting the use of whole genome amplification for genomic analysis.

Gréen, HenrikLinköpings universitet,KTH,Genteknologi,Science for Life Laboratory, SciLifeLab,Avdelningen för läkemedelsforskning,Hälsouniversitetet,Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden(Swepub:liu)hengr89 (författare)
Solnestam, Beata WerneKTH,Genteknologi,Science for Life Laboratory, SciLifeLab,Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden(Swepub:kth)u1n13z7k (författare)
Zajac, PawelLaboratory for Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden (författare)
Huss, MikaelStockholms universitet,Institutionen för biokemi och biofysik(Swepub:su)mihu5153 (författare)
Orear, CedricGenomics Unit, Institut Gustave Roussy, Villejuif, France (författare)
Validire, PierreDepartment of Pathology, Institut Mutualiste Montsouris, Paris, France (författare)
Bjursell, MagnusAstraZeneca R&D, Mölndal, Sweden (författare)
Lundeberg, JoakimKTH,Genteknologi,Science for Life Laboratory, SciLifeLab,Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, Stockholm, Sweden(Swepub:kth)u1qkn9kw (författare)
KTHGenteknologi (creator_code:org_t)

Ingår i:Biochemical and Biophysical Research Communications - BBRC: Elsevier BV425:2, s. 379-3830006-291X1090-2104

Av författaren/redakt...: Hasmats, Johanna; Gréen, Henrik; Solnestam, Beata ...; Zajac, Pawel; Huss, Mikael; Orear, Cedric; visa fler...; Validire, Pierre; Bjursell, Magnus; Lundeberg, Joaki ...; visa färre...

Om ämnet

Av lärosätet: Kungliga Tekniska Högskolan; Karolinska Institutet; Linköpings universitet; Stockholms universitet

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