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Efficient characterization of retro-, lenti-, and foamyvector-transduced cell populations by high-accuracy insertion site sequencing

In English

Författare

  • M Schmidt
  • H Glimm
  • M Wissler
  • G Hoffmann
  • Karin Olsson
  • S Sellers
  • D Carbonaro
  • JF Tisdale
  • C Leurs
  • H Hanenberg
  • CE Dunbar
  • HP Kiem
  • Stefan Karlsson
  • DB Kohn
  • D Williams
  • C von Kalle
Visa allt

Summary, in English

The identification of unknown genomic flanking DNA sequences can be used for the molecular monitoring of retro-, lenti- and foamyviral integration, transgenes in early embryogenesis, insertional mutagenesis, cell fate, and stem cell plasticity. Most existing methods reflect shortcomings in sensitivity and or specificity, thus limiting genomic sequencing of unknown flanking DNA to clonal preparations. The application of linear amplification-mediated PCR (LAM-PCR), a recently developed direct sequencing technique for flanking DNA, should circumvent current limitations in different research fields. This technique combines preamplification of target DNA with a unique succession of enzymatic reactions on solid-phase. Using LAM-PCR, we show the previously unfeasible in vivo retro-, lenti- and foamyvirus integration site analysis in primate peripheral blood hematopoietic cells and human xenograft hematopoiesis. In light of two severe adverse events that occurred in a clinical SCID-X1 gene therapy trial, in vivo monitoring of the reinfused transduced cell pool by integration site analysis will be an important component of each gene transfer and therapy study aimed at clinical use.

Publiceringsår

2003

Språk

Engelska

Sidor

112-121

Publikation/Tidskrift/Serie

Annals of the New York Academy of Sciences (HEMATOPOIETIC STEM CELLS 2002: GENETICS AND FUNCTION: Fourth International Symposium)

Volym

996

Dokumenttyp

Konferensbidrag

Förlag

New York Academy of Sciences

Ämne

  • Hematology

Nyckelord

  • proviruses
  • therapy
  • insertion sequence elements
  • investigative techniques
  • gene
  • stem cells

Conference name

HEMATOPOIETIC STEM CELLS 2002: GENETICS AND FUNCTION: Fourth International Symposium

Conference date

0001-01-02

Status

Published

ISBN/ISSN/Övrigt

  • ISSN: 0077-8923