The segregating neutral sites should be interpreted as a sample of neutral marker segregating sites.
Our findings suggest that the initial mass selection used to form the base population of the maritime pine breeding program was not only successful in terms of the initiation of a program to develop improved varieties [ 58], but also efficient for the sampling of neutral genetic diversity from the Landes forest.
The approach described in this study can be used, in general, to considerably improve the efficiency of RPLC method development, particularly for neutral samples.
Samples were taken periodically for neutral lipids analysis.
A sub-sample of neutral markers was thus defined and used for the genetic diversity analysis.
In a typical experiment, DNA samples are enzymatically hydrolyzed to free deoxyribonucleosides with the resulting sample analyzed by LC-MSn for neutral loss of m/ z = 116 corresponding to the deoxyribose group.
The monosaccharide concentrations were then quantified by HPAEC-PAD (Thermo Fisher Scientific Inc., Bannockburn, IL, USA): for neutral sugars, a sample size of 25 μL was injected onto a Dionex CarboPac PA20 column (3 × 30 mm guard and 3 × 150 mm analytical) and eluted at 30°C using an isocratic mobile phase of 18 mM KOH at 0.4 mL/minute over 11 minutes.
Resource constraints precluded us from performing all analyses on every sample, so we selected 15 sample pairs for both neutral and acid extractions, 25 for neutral extractions only, and 5 for acid extractions only (for a total of 40 sample pairs extracted for neutrals and 20 for acids).
Samples were analyzed by wet chemistry for neutral detergent fiber (NDF), according to procedures by Van Soest et al. (1991).
However, a parallel sample run with no SGFRs added showed zero reads mapping to the regions of select fused gene junctions, and therefore the COLO829 can be considered to be a high complexity neutral background sample for this study.
