Using EF5 as a hypoxia marker, we determined that the spheroids used in these experiments contained hypoxic regions (Koch, 2002).
In order to place a mutation in the genetic interval between the closest marker and another linked marker we determined whether recombinations for both of them were correlated.
Using the ribulose 6-phosphate carboxylase oxygenase (rbcL) coding region as a marker, we determined species composition at these sites and compared these data to a typical endolithic population that was previously described from Pisciarelli (PBen site, Fig. 1B) [ 6].
Based on the combination of the eight markers, we determined the final genotype using an arbitrary implementation.
Using strains co-expressing Golgi markers, we determined that GeaA1-GFP partially co-localizes with the early Golgi marker SedV, although to a lesser extent than GeaA-GFP (Fig. 4B and D).
For all individual aligned orthologous markers, we determined the amino acid and nucleotide substitution models that best fit the data using the command-line mode of ProtTest v1.4 [ 34] and ModelGenerator v0.85 [ 35], respectively.
Similarly, for the other Type III markers, we determined that four markers (No. 1, 9, 13 and 15) could be assigned to three chromosomes, five markers (No. 2, 4, 5, 11 and 23) to two chromosomes, and two markers (No. 17 and 20) to a single chromosome (Table 2).
In subsequent studies where we stained back to back sections with antibodies against the cytokines IL-1β, IL-6 and IL-8 and with leukocyte cell surface markers we determined that protein expression of these pro-inflammatory cytokines was largely (although not exclusively) confined to invading leukocytes [ 4].
Using a number of randomly selected loci genotyped in the Cacao6kSNP array as a pool of markers, we determined the probability of successfully differentiating accessions with 10%, 16.2% (as observed with UF273 Type I and Type II) and 30% genetic variation (Fig. 5).
After acquiring a statistic of genetic incompatibility for each pair of markers, we determine statistical significance using a familywise 5% type I error rate (discussed later).
To identify robust, polymorphic markers for population studies we determined the level of SSR polymorphism in a small collection of six Brachypodium lines (Bd1-1, Bd3-1, Bd21-3Bd18-1, Bd21, Bd21-3) using 621 primer pairs (144 from ESTs and 477 from BES) (additional file 1).
