Protocol

Transduction of Primary Hematopoietic Cells by Retroviral Vectors

This protocol was adapted from "Retroviral Vectors," Chapter 2, in Gene Transfer: Delivery and Expression of DNA and RNA (eds. Friedmann and Rossi). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2007.

INTRODUCTION

Noncycling cells are relatively resistant to transduction with retroviral vectors. Because most immortalized cell lines are actively proliferating, this is not an issue. However, for many primary cells, especially quiescent populations such as primitive progenitor and stem cells, the gene-transfer rate can be particularly low. This protocol describes transduction of primary hematopoietic cells. Two interventions are combined to maximize gene transfer in hematopoietic progenitor cells: (1) cytokines and other growth factors are used to stimulate cell cycling in hematopoietic cells, and (2) matrix proteins such as fibronectin are used to mediate colocalization of target cells and vector.