Defense mechanisms against herbivory: expression regulation of spruce gene family members representative of the phenylpropanoid pathway

A 21.8k spruce EST-spotted microarray was utilized to examine the genetic basis of constitutive defenses against herbivory. From twelve-year-old trees originating from partial diallel pedigree that segregates for resistance against Pissodes strobi, bark tissue of apical leaders was harvested at the onset of seasonal growth and natural weevil activity in the field. Our experimental design is based on a priori known genotype. We identified cross 26 from ♀PG87*♂PG165, cross 27 from ♀PG87*♂PG117, cross 29 from ♀PG21*♂PG165 and cross 32 from ♀PG21*♂PG117, respectively, forming a factorial diallel which showed widest segregation, based on a higher number of segregating SNP loci with genotypes significantly different between the most extreme resistance categories. The distant pair design introduced by ({Fu, 2006}) was modified for outbreds and maximized direct comparisons between different alleles at each locus. Our design resulted in 94 hybridisations profiling 48 individuals in cross 26, 36 in cross 27, and 50 in cross 29 as well as 54 individuals in cross 32. One criterion for identifying positional candidate genes involved that any association between trait variation and genotype variation had to be detected within 10 cM map distances; positional candidate genes for the studied phenotypic trait were then identified by strong collocations of transcript abundance variation and overlap of at least 40% of their expression quantitative trait loci (eQTLs) with phenotypic trait QTLs. 10,000 randomizations were run (p < 0.05). The experimental design optimized genetic distances between probe pairs on 94 two-color microarrays to profile 188 interior spruce F1-hybrids, independently grown and harvested.