GSM3003718: Pooled RNA Ccnb3 MO 16-cell 3; Ciona intestinalis; RNA-Seq
For single cells, cDNA was prepared directly from cell lysate. For pooled embryos total RNA was purified using Trizol. cDNA was synthesized and amplified from the poly-A containing RNA transcripts in the single-cell lysate using the Smart-Seq2 method. cDNA from cells at 4-cell stage was amplified by 15 cycles of PCR, while 16 cycles of PCR was applied on the cDNA from cells of 8-cell and 16-cell stage embryos. The amplified cDNA samples were cleaned up with Ampure XP beads (Beckman Coulter, CA, USA), quantified by Qubit fluorometer (Invitrogen, CA, USA), and examined on Bioanalyzer with High Sensitivity DNA chips (Agilent, CA, USA) for size distribution. Nextera DNA library prep kit (Illumina, CA, USA) was used to convert the cDNA samples to Illumina sequencing libraries. For pooled embryo samples, cDNA synthesis and sequencing was performed as described for single-cells with the exception that 12 cycles of PCR were performed.
GEO Accession
GSM3003718