GSM8001136: 4-cell embryo, heat-exposed sperm fertilized, rep 9; Mus musculus; RNA-Seq
RNA was isolated using RNA-Clean XP beads Illumina Nextera XT Kit Sperm pellets were resuspended in lysis buffer supplemented with 33 mM DTT and 0.66 mg/mL Proteinase K and incubated at 60°C for 15 min. After incubation, an equal volume of water and TRIzol was added to each sample. The samples were transferred to phase lock tubes for phase separation and precipitation. Small RNA was purified using denaturing polyacrylamide gel electrophoresis. Single embryos were collected into 1 x TCL buffer with 1% beta-mercaptoethanol and stored at -80°C prior to sequencing. At time of sequencing embryos were further lysed in fresh lysis buffer and RNA was captured using RNA Clean XP beads. For sperm samples - Illumina Tru-seq Small RNA Library Preparation Kit and single embryo samples - Nextera XT kit (Illumina)