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In each cord a minimum of 12 sections was used for each marker.
Consistent exposure parameters were used for each marker to allow the direct comparison of fluorescent intensity across experimental conditions.
All microscopy done on live animals was performed on a Zeiss Axioplan2 microscope, with a consistent exposure time used for each marker assayed.
25 ng of genomic DNA was used for each marker.
Details of the antibodies and techniques used for each marker are shown in Table 1.
The Wald statistical test was used for each marker to test the null hypothesis that the marker's effect was zero.
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We performed this analysis on 17 unrelated families from the breeding population, using for each marker the allele frequency (p) estimated on the Aquitaine G0 genotyping dataset (212 samples).
One representative section on every 10th slide corresponding to every 500 µm along the cord was used for each cell marker.
3 embryos from each group (control or mutant) were used for each molecular marker labeling, and 2 wild-type embryos from each developmental stage (E6.5 or E7.5) were used for in situ hybridization of Pik3c3.
Table 2 summarizes antigens, primary antibodies, clone, source, optimal dilution, and scoring system used for each immunohistochemical marker.
PCR primers and cycling conditions used for each of the markers are referenced in [ 27], with information on additional LSU and COI primer derivation as follows.
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