Collagen transcript analysis by qPCR vs. RNA-seq.

<p>Total RNA was isolated from whole day 13 conceptuses, while for day 19, 21, and 25 conceptuses, yolk sac fluid was carefully removed and heart and blood vessel structures were separated from extraembryonic membranes. The extraembryonic membrane samples consisted of yolk sac and chorion on day 19 and 21 whereas on day 25, the samples contained yolk sac and allantochorion membranes. Collagen transcripts analyzed were <i>COL4A1</i>, <i>COL4A5</i>, <i>COL4A6</i>, <i>COL5A2</i>, <i>COL6A1</i>, <i>COL6A2</i>, and <i>COL6A3</i>. Gray bars indicate data from RNA-seq analysis, and dark bars represent data from qPCR analyses. Note that RNA-seq data represent transcripts extracted from day 19, 21 and 25 trophectoderm and/or chorionic membrane, whereas RNA from days 13, 19, 21 and 25 were analyzed by qPCR. An average of at least three conceptuses was analyzed for each time point, and the SEM is shown. Please note that values on Y-axis resulted from the generation of a standard curve for each transcript. In doing so, the highest amount in each transcript was treated as “1”, and relative values were presented.