Measurements of the expression of the gene DEAD
(Asp-Glu-Ala-Asp) box polypeptide 4 (DDX4 or VASA)
Mm00802445_m1 in the neonate and adult SSCs were analyzed
with Dynamic Array chips (Fluidigm). A housekeeping
gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH)
Mm99999915_g1 was used for normalization, in different types
of cultured cells. SSCs were picked with a micromanipulator,
lysed with a lysis buffer solution containing 9 μl RT-PreAmp
Master Mix (5.0 μl Cells Direct 2× Reaction Mix) (Invitrogen,
USA), 2.5 μl 0.2× assay pool, 0.2 μl RT/Taq Superscript III
(Invitrogen, USA) and 1.3 μl TE buffer. The number of RNAtargeted
transcripts was measured using TaqMan PCR assays
on the BioMark Real-Time quantitative PCR (qPCR) system.
Each sample was analyzed in two technical repeats. The Ct
values were examined using GenEx software from MultiD for
analysis (4 (link), 5 (link)).
(Asp-Glu-Ala-Asp) box polypeptide 4 (DDX4 or VASA)
Mm00802445_m1 in the neonate and adult SSCs were analyzed
with Dynamic Array chips (Fluidigm). A housekeeping
gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH)
Mm99999915_g1 was used for normalization, in different types
of cultured cells. SSCs were picked with a micromanipulator,
lysed with a lysis buffer solution containing 9 μl RT-PreAmp
Master Mix (5.0 μl Cells Direct 2× Reaction Mix) (Invitrogen,
USA), 2.5 μl 0.2× assay pool, 0.2 μl RT/Taq Superscript III
(Invitrogen, USA) and 1.3 μl TE buffer. The number of RNAtargeted
transcripts was measured using TaqMan PCR assays
on the BioMark Real-Time quantitative PCR (qPCR) system.
Each sample was analyzed in two technical repeats. The Ct
values were examined using GenEx software from MultiD for
analysis (4 (link), 5 (link)).
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