Dosage and stability tests of antisense morpholinos in mouse embryos. (A) The developmental potential of embryos was evaluated using 1-20 mM doses of control nonsense MO, which was conjugated to FITC fluorescent dye and microinjected into the cytoplasm of pronuclear-stage embryos. Upper panel: Phase contrast images. Lower panel: Fluorescence images. >Scale bar: 30 μm. (B) Granzyme G mRNA blocking efficiency test following microinjection of different doses of granzyme G-specific antisense MO (0, 0.2, and 2 mM) into 1-cell stage embryos. Total RNA was isolated from 2-cell stage experimental embryos, followed by RNase III digestion, purification, and subjection to RT-PCR analysis. In the control panel, mRNA extracted from the control MO-injected embryos was used as an un-blocked control (lane 5), mRNA from pregnant mouse uterine GMG cells was used as a positive control (Pos; lane 6), and reverse transcriptase-omitted mRNA template was used as a negative control (Neg; lane 7). The results are representative of three experiments.
Tsai et al. BMC Developmental Biology 2010 10:88 doi:10.1186/1471-213X-10-88