A Comparative Analysis of Library Prep Approaches for Sequencing Low Input Translatome Samples

We evaluated the performance of 5 library prep protocols by using total mRNA and IP RNA of mouse liver,we found all the 5 library preparation kits detect more enrichment effects than depletion effect. The profiles being generated by SMARTer kit is different than all other kits. Cell-type specific RNA from livers of P30 mice was obtained using the RiboTag protocol. Briefly, liver tissue expressing HA-tagged ribosomes in specific cells was homogenized followed by immunoprecipitation of the tagged ribosomes using an HA antibody. The RNA was extracted from the immunoprecipitated ribosomes to obtain cell-type specific enriched transcripts (IP samples). An aliquot of the homogenized tissue before immunoprecipitation was used for total RNA extraction (input samples).