Lineage specification of Flk-1+ progenitors is associated with divergent Sox7 expression in cardiopoiesis
DOI: 10.1016/j.diff.2008.10.012
Authors: Timothy J. Nelson; Anca Chiriac; Randolph S. Faustino; Ruben J. Crespo-Diaz; Atta Behfar; Andre Terzic
Step 1: iPSC/ESC maintainance
day 0 ~ 1 (duration missing in database; assumed 1 day)
culture “unspecified method” mouse ESC in “GMEM”. Medium should contain factors shown in the following table.
| concentration | unit | |
|---|---|---|
| pyruvate | — | — |
| NEAA | — | — |
| 2-mercaptoethanol | — | — |
| FCS | 7.5 | % |
| LIF | — | — |
Step 2: Primitive streak 1
day 1 ~ 3
culture “spheroid” mouse ESC in “GMEM” for “2” days. Medium should contain factors shown in the following table, and should be changed once in “2” days.
| concentration | unit | |
|---|---|---|
| pyruvate | — | — |
| NEAA | — | — |
| 2-mercaptoethanol | — | — |
| FCS | 20 | % |
| TNF-a | 3.0e-5 | g/l |
Ideally, “dummy” should express “POU5F1”, “Fgf4” at the end of culture.
Step 3: mesoderm 1
day 3 ~ 6
culture “spheroid” mouse ESC in “GMEM” for “3” days. Medium should contain factors shown in the following table, and should be changed once in “3” days.
| concentration | unit | |
|---|---|---|
| pyruvate | — | — |
| NEAA | — | — |
| 2-mercaptoethanol | — | — |
| FCS | 20 | % |
Ideally, “dummy” should express “KDR”, “CXCR4”, “Sox1”, “Sox7”, “SOX17”, “Gsc”, “Lhx1” at the end of culture.
Step 4: mesoderm 2
day 6 ~ 7 (duration missing in database; assumed 1 day)
culture “adherent” mouse ESC in “unspecified medium”. For ECM, use “Gelatin” at “N/A” “N/A”.
Ideally, “dummy” should express “Nkx2.5”, “Mef2c”, “Myocd”, “GATA4” at the end of culture.