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

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

Forskningsoutput: Kapitel i bok/rapport/Conference proceedingKonferenspaper i proceedingPeer review

Sammanfattning

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.
Originalspråkengelska
Titel på värdpublikationAnnals of the New York Academy of Sciences (HEMATOPOIETIC STEM CELLS 2002: GENETICS AND FUNCTION: Fourth International Symposium)
FörlagNew York Academy of Sciences
Sidor112-121
Volym996
StatusPublished - 2003
EvenemangHEMATOPOIETIC STEM CELLS 2002: GENETICS AND FUNCTION: Fourth International Symposium -
Varaktighet: 0001 jan. 2 → …

Publikationsserier

Namn
Volym996
ISSN (tryckt)0077-8923

Konferens

KonferensHEMATOPOIETIC STEM CELLS 2002: GENETICS AND FUNCTION: Fourth International Symposium
Period0001/01/02 → …

Ämnesklassifikation (UKÄ)

  • Hematologi

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