BacMam-mediated gene transfer into neural cell types

Abstract

Modified baculoviruses containing a mammalian promoter, known as BacMam, have emerged as a key method for labeling diverse cell types, owing to its high transduction efficiencies, minimal cytopathic effects and the ability to carry DNA fragments larger than 40 KB. The transient expression without viral replication or integration into the mammalian genome makes BacMam an ideal method to label primary and adult stem cells. In the current study, we describe how BacMam enables efficient transduction and robust gene expression in neural stem cells (NSC) and progenitor cells. Utilizing the H9 human embryonic stem cell (hESC), we generated neurospheres, neural rosettes and Neural Stem Cells (NSC) and transduced them with BacMam 2.0 GFP. Commercially available human NSCs (h-NSC) and rat fetal NSCs (rNSCs) were also transduced with BacMam 2.0 GFP. Primary rat neurons and post-differentiated human NSCs were transduced with either BacMam 2.0 GFP or Plasma Membrane-GFP BacMam 2.0 and Synaptophysin-RFP BacMam 2.0 to assess the capability of gene expression in mature neural cell types. Using a GFP marker, we demonstrated that the BacMam virus can consistently transduce neural stem cells with high efficiency and low toxicity. Transduction was also enabled in differentiated NSC, and the expression in primary neurons was dependent on the age of the culture, with preferential labeling of astrocytes in older cultures. BacMam mediated gene delivery provides a superior platform for introducing diverse gene elements, creating assay-ready cells among young neuronal cells and mature astrocytes.