Transcriptional analysis of murine cortical astrocytes with adeno-associated viral transduction with and without 4x6T miR cassette, vs transgenically labeled astrocytes
We generated adeno-associated Cre viral vectors with improved astrocyte-specific expression by including a cassette of miRNA targeting sequences (4x6T) to decrease off-target expression. We delivered these viruses in Ribotag mice using the PHP.eB serotype, either intracortically or systemically. We compared these samples with mice that received intracortical AAV2/5::GfaABC1D-Cre and transgenic Aldh1l1-CreERT2/Ribotag mice. We sequenced whole-tissue RNA as well as ribosomally-loaded astrocyte-enriched mRNA to assess cell-type specificity of viruses as well as virus-related transcriptomic changes. Overall design: Four cohorts were generated: Aldh1l1-CreERT2/Ribotag transgenic mice (control mice with highly astrocyte-specific labeled ribosomes); AAV2/5::GfaABC1D-Cre intracortical injection mice (standard astrocyte-enriched virus); PHP.eB::GfaABC1D-Cre-4x6T intracortical injection mice; and PHP.eB::GfaABC1D-Cre-4x6T systemic injection mice. Each cohort included 4 mice, and each mouse had both input (whole-cortex) and immunoprecipitated (ribosomally loaded, astrocyte-enriched) mRNA samples.