Fig. 3. The different BMP inhibitory activities between TSKA and TSKB. TSKA (A) or TSKB (B) mRNA was injected into a ventral vegetal blastomere at the eight-cell stage and the embryos were developed until stage 31. Arrow indicates a secondary axis. (C) Western blot analysis of TSKA-Myc-His, TSKB-Myc-His and BMP4-Flag proteins used. (D) Co-immunoprecipitation of TSKA-Myc-His or TSKB-Myc-His and BMP4-Flag. After immunoprecipitation with nickel-chelating resins, the complexes were washed under low or high stringency conditions. (E) Co-immunoprecipitation of TSKA-Myc-His or TSKB-Myc-His and BMP4-Flag with different amounts of BMP4. The numbers above the column show the volume (µl) of COS-7 cell supernatant, including BMP4 in the total volume (1 ml). (F) Co-immunoprecipitation of TSKB-Myc-His and BMP7-Flag. (G,J) Transplantation schemes. (H,I) TSKA- or TSKB-producing cells (orange in G) were grafted together with the middle primitive streak (MPS; green in G) on the right-hand side. The mock-transfected COS-7 cells (white in G) were grafted with MPS on the left-hand side. (K,L) TSKA or TSKB-producing cells (orange in J) were grafted together with cells expressing VG1 (purple in J) and WNT1 (blue in J) on the right-hand side. The mock-transfected COS-7 cells (white) were grafted with cells expressing VG1 and WNT1 on the left-hand side.