A recombinant retrovirus encoding alkaline phosphatase confirms clonal boundary assignment in lineage analysis of murine retina.

Recombinant retroviruses encoding the histochemically detectable enzyme beta-galactosidase have been used to investigate lineage in the vertebrate nervous system. Identification of the descendants of individual progenitors is straightforward when progeny cells are arranged in a reproducible, clustered pattern, but difficulties in interpretation arise when progeny migrate extensively and/or in an irregular pattern. To better resolve clonal boundaries, additional histochemical marker viruses that engender distinctive reaction products can be used in combination with lacZ-bearing viruses. To this end, we have created a retrovirus vector, DAP, encoding an easily assayable enzyme, human placental alkaline phosphatase. DAP was found to be at least as useful as a lacZ-encoding retrovirus (e.g., BAG) with respect to high viral titer, stability of expression, and in identification of infected cells in vivo. Moreover, it was found to be neutral with respect to postnatal rodent retinal development and offered superior staining characteristics relative to lacZ. Coinfection of rodent retina with DAP and BAG allowed an examination of the clonal nature of radial arrays of labeled retinal cells that previously had been described as products of a single infected progenitor. Of 1100 radial arrays examined for the presence of both DAP- and BAG-infected cells, only 1.2% were the result of infection with more than one virus.