Fig. 4. Ascl1 is required for neurogenic potential in animal cells. (A) qPCR comparison of mesendoderm gene expression (means±s.d.) in vegt-injected animal cap explants at sibling stage 10.5. Control and Ascl1 MOs (60 ng) were injected at one-cell stage. vegt mRNA (100 pg) was injected into animal pole at two-cell stage and animal caps were dissected at stage 8.5 and cultured to the sibling stage 10.5 for RT-qPCR analysis. **P<0.01. (B) Animal caps at the sibling stage 15 in situ hybridized with tubb2b. Control MO (cMO) and Ascl1 MOs (AMOs) (60 ng) were injected at one-cell stage, fgf8a mRNA (100 pg) was injected into animal pole at two-cell stage, and animal caps were dissected at stage 8.5 and cultured to the sibling stage 15 followed by in situ hybridization. (C) qPCR comparison of the expression of tubb2b (means±s.d.) in animal cap explants. The injection and dissection schemes were the same as described in B, except that the RT-qPCR analysis was performed at the sibling stage 18. (D) Animal cap explants injected with Ascl1 MOa+b or spMO isolated at stage 9 treated with or without Noggin protein (350 ng/ml) for 4 h, and then cultured to sibling stage 18 followed by qPCR detection of sox2 expression (means±s.d.). **P<0.01. (E) RT-PCR verification of injection of Ascl1a spMO blocked ascl1a splice in stage 10.5 embryos. PCR using genomic DNA template also included as a control for the size of product amplified from unspliced mRNA/cDNA templates. (F) qPCR examination of gene expression in Ascl1a spMO at stage 10.5 (means±s.d.).
Image published in: Gao L et al. (2016)
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