GSM8683134: mOocyte, WtFloxCre, rep4; Mus musculus; RNA-Seq
The isolated full-grown oocytes at the germinal vesicle (GV) stage were washed in washing medium [0.1% PBA in PBS] and collected manually with 0.5 μL of the washing medium in 2.125 μl cell lysis buffer [10X Lysis Buffer (TAKARA), RNase Inhibitor (TAKARA), and RNase-free water (Nacalai)] in 0.2 mL 8-strip tubes. Three oocytes were collected in each tube with one or two replicates from each mouse. The SMART-seq v4 Ultra Low Input RNA Kit for Sequencing (Takara) was used to reverse transcribe poly(A) RNA and amplify full-length cDNA. Samples were amplified for 13 cycles in 8-strip tubes. After the cDNA was adjusted to 0.5 ng in 1.25 μL of elution buffer [10mM Tris-HCl, pH8.5], Tn5 tagmentation-based reaction was performed with 1/4 volume of the Nextera XT DNA Library Preparation Kit (Illumina) with Nextera XT Index Kit (Illumina) according to manufacturer's protocol. Library DNA was amplified with 12 cycles of PCR and purified using 1.8× volume of SPRISelect (Beckman Coulter) and eluted into 12 μL of the elution buffer [10mM Tris-HCl, pH8.5].