Transcriptomic profiling of adult zebrafish photoreceptors

Identifying the transcription factors required to specify photoreceptor subtypes is critical to understand the normal development of the retina and to inform cell-replacement therapies to restore vision. RNA-seq is a powerful way to identify novel genes expressed in particular cell subtypes. Although RNA-seq approaches have been used to identify genes differentially expressed between photoreceptor subtypes in many species, the limited transcriptome depth derived from single-cell techniques constitutes a barrier in the reliable detection of transcription factors. To obtain a deep, high-quality RNA-seq dataset from zebrafish photoreceptors, we manually collected pools of photoreceptors of a single subtype. Overall design: We identified photoreceptors using well-characterized transgenic lines that express fluorescent proteins in each subtype with high specificity, including rods—Tg(xOPS:GFP), UV cones—Tg(opn1sw1:GFP), S cones—Tg(opn1sw2:GFP), M cones—Tg(opn1mw2:GFP) and L cones—Tg(thrb:tdTomato). We manually collected pools of 20 dissociated but healthy photoreceptors of a single subtype for each of our samples, while actively avoiding cellular debris and other contaminants.