(A) mRNA expression of vinculin, paxillin, FAK, and Tensin1 was assessed by qRT-PCR in yolk sacs harvested at 9.0 dpc (15-20 somite stage) versus 9.5 dpc (25-30 somite stage). Relative to corresponding WT yolk sacs isolated from littermates at either 9.0 dpc or 9.5 dpc, mRNA expression of these genes is increased in Raldh2−/− yolk sacs specifically at 9.5 dpc and not prior at 9.0 dpc. Transcript levels were normalized by using β-actin as the internal control. (B) Protein levels of FAK, paxillin, and vinculin were determined by Western blot in 8.0 dpc WT, Raldh2 heterozygote (+/−), and Raldh2 null (−/−) yolk sacs isolated at the 3-6 somite stage prior to cardiac function and establishment of blood flow. At this stage, protein levels in Raldh2 −/− yolk sacs were not found to be different from WT or Raldh2 +/− yolk sacs. Actin was used as the loading control.