Aplysia kinesin cargo analysis

Subcellular RNA targeting was performed by immunoprecipitation with the Aplysia kinesin antibody followed by high throughput sequencing of the cargo bound to kinesin. All bound RNAs were extracted and sequenced. Specifically, we developed a reduced representation sequencing method that can detect low abundant transcripts. Our method greatly reduces the amount of sequencing with more accurate quantification. Basically, sequential addition of adaptors paired with controlled digestion of cDNA generates a one transcript on read type of analysis. Combined 5’ with 3’ targeted libraries produces greater coverage of individual transcripts and high quality assembly. Our initial analysis allowed us to identify several thousands of neuronal transcripts bound to kinesis cargo including a diversity of receptors and transcription factors essential for the establishing of cell identity and plasticity.