GENOME TO PHENOME VIA SINGLE-CELL ALTERNATIVE TRANSCRIPTOME ANALYSIS IN CATTLE

The overall goal of this project is to understand how cellular heterogeneity bridges the static genome to the dynamic phenome in general. Specifically, we will adapt our whole transcriptome start and termini site sequencing (WTSS-seq and WTTS-seq) methods into single cell RNA sequencing (scRNA-seq) methods to facilitate functional annotation of bovine genomes. These novel tools will help broad research communities and industry partners to advance their animal genome research programs and their applications by focusing on the Genome to Phenome in the next 10 years and beyond.Specific Objective 1: Develop a single cell WTSS-seq (scWTSS-seq) method to study cellular heterogeneity in use of alternative transcription start (ATS) sites. Our scWTSS-seq method will entail six major steps: rRNA blocking, reverse transcription, RNA removal, PCR amplification, size selection and library sequencing.Specific Objective 2: Develop a single cell WTTS-seq (scWTTS-seq) method to study cellular heterogeneity in use of alternative polyadenylation (APA) sites. Our scWTTS-seq method will be completed with six major steps: reverse transcription, RNA removal, second-strand cDNA synthesis, PCR amplification, size selection and library sequencing.