Identification of tens of thousands of novel gene loci in the human and mouse genomes through targeted full-length long-read RNA sequencing
Identification of tens of thousands of novel gene loci in the human and mouse genomes through targeted full-length long-read RNA sequencing
Mouse samples were obtained from the PRBB (Barcelona Biomedical Research Park) Animal Facility. The samples were collected from both adult and embryonic mice of the C57BL/6 strain. Adult mice were selected based on standard laboratory guidelines, and embryos were harvested at the desired developmental stages under sterile conditions. All procedures were performed in accordance with institutional animal care and ethical guidelines to ensure the welfare of the animals. Total RNA Extraction Protocol: Total RNA samples were extracted from tissues or cell lines using the Thermo Fisher PureLink RNA Mini Kit. This kit utilizes a silica membrane-based purification system, which allows for the efficient isolation of high-quality RNA by binding nucleic acids to a silica membrane. Samples are lysed (using Trizol) and homogenized, and RNA is selectively bound to the membrane while impurities are washed away. The final purified RNA is eluted in RNase-free water, ensuring it is suitable for downstream applications. SMARTerStranded: The SMARTer Stranded RNA-Seq protocol is a next-generation sequencing method designed for generating high-quality, strand-specific RNA libraries for Illumina short-reads. It preserves strand information by incorporating adapters to maintain the orientation of RNA transcripts.