The first step toward generating whale organoids is to dedifferentiate fibroblasts into reprogrammable induced pluripotent stem cells (iPSCs). To accomplish this step, we must transfect the fibroblasts with episomal vectors containing the 5 reprogramming vectors. The transfection process creates small tears in the cell membranes which allow the episomal vectors to enter the cells.

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We have attempted two transfections thus far on human cells and one transfection with bottlenose dolphin cells. Our main challenge has been obtaining a starting cell count high enough that we have a sufficient number of cells that survive transfection and end up with all five reprogramming vectors incorporated. 

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To continue moving towards accomplishing this goal, we will re-attempt transfection with human cells and plan to increase our starting cell count by an order of magnitude. We will use this transfection to continue to optimize the timeline of passaging and media changes in preparation for conducting a transfection on Cuvier's beaked whale and other whale and dolphin cells. 

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For more background on the process of creating heart organoids, see Katherine Krieger's blog post here.