GeneticsRandom amplified polymorphic DNA markers tightly linked to a gene for resistance to white pine blister rust in sugar pine ( ABSTRACTWe have genetically mapped a gene for resistance to white pine blister rust (Cronartium ribicola Fisch.) in sugar pine (Pinus lambertiana Dougl.) by using an approach which relies on three factors: (i) the ability to assay for genetic markers in the haploid stage of the host's life cycle, using megagametophyte seed tissue; (ii) a simple and clearly defined pathosystem; and (iii) the use of random amplified polymorphic DNA (RAPD) markers that can be quickly and efficiently evaluated. Resistance to white pine blister rust in sugar pine is known to be controlled by a single dominant gene (R). Maternal segregation ofR and dominant RAPD markers were scored simultaneously following collection of megagametophytes for DNA assays and seedling inoculation with C. ribicola. Bulked samples of haploid megagametophyte DNA from resistant and susceptible offspring of segregating full-sib and half-sib families were used to evaluate 800 random decanucleotide primers. Ten loci linked with the gene for resistance to white pine blister rust were identified and segregation data were obtained from five families. Six of the linked markers were within 5 centimorgans of the gene, and one marker was 0.9 centimorgan from R. These and other markers derived by this approach may provide starting points for map-based cloning of this important gene.

Sugar pine (Pinus lambertiana Douglas) is within the subgenus Strobus with an estimated genome size of 31 Gbp. Transcriptomic resources are of particular interest in conifers due to the challenges presented in their megagenomes for gene identification. In this study, we present the first comprehensive survey of the P. lambertiana transcriptome through deep sequencing of a variety of tissue types to generate more than 2.5 billion short reads. Third generation, long reads generated through PacBio Iso-Seq have been included for the first time in conifers to combat the challenges associated with de novo transcriptome assembly. A technology comparison is provided here to contribute to the otherwise scarce comparisons of second and third generation transcriptome sequencing approaches in plant species. In addition, the transcriptome reference was essential for gene model identification and quality assessment in the parallel project responsible for sequencing and assembly of the entire genome. In this study, the transcriptomic data were also used to address questions surrounding lineage-specific Dicer-like proteins in conifers. These proteins play a role in the control of transposable element proliferation and the related genome expansion in conifers.

The majority of genomic research in conifers has been conducted in the Pinus subgenus Pinus mostly due to the high economic importance of the species within this taxon. Genetic maps have been constructed for several of these pines and comparative mapping analyses have consistently revealed notable synteny. In contrast, little genomic research has been conducted on the Pinus subgenus Strobus, even though these pines have strong ecological relevance. We report a consensus genetic linkage map for sugar pine (Pinus lambertiana Dougl.) constructed with 399 single nucleotide polymorphisms markers derived from annotated genes. The map is 1,231 cM in length and organized into 19 linkage groups. Two of the mapping populations were derived from trees that were segregating for the major gene of resistance (Cr1) to Cronartium ribicola, the fungal pathogen responsible for white pine blister rust. The third mapping population was derived from a full-sib cross segregating for partial resistance to white pine blister rust. In addition, we report the first comparative mapping study between subgenera Strobus and Pinus. Sixty mapped markers were found in common between sugar pine and the loblolly pine reference map with 56 of them (93%) showing collinearity. All 19 linkage groups of the sugar pine consensus map coaligned to the 12 linkage groups of the loblolly pine reference map. The syntenic relationship observed between these two clades of pines provides a foundation for advancing genomic research and genetic resources in subgenus Strobus.

Conifers are the dominant plant species throughout the high latitude boreal forests as well as some lower latitude temperate forests of North America, Europe, and Asia. As such, they play an integral economic and ecological role across much of the world. This study focused on the characterization of needle transcriptomes from four ecologically important and understudied North American white pines within the Pinus subgenus Strobus. The populations of many Strobus species are challenged by native and introduced pathogens, native insects, and abiotic factors. RNA from the needles of western white pine (Pinus monticola), limber pine (Pinus flexilis), whitebark pine (Pinus albicaulis), and sugar pine (Pinus lambertiana) was sampled, Illumina short read sequenced, and de novo assembled. The assembled transcripts and their subsequent structural and functional annotations were processed through custom pipelines to contend with the challenges of non-model organism transcriptome validation. Orthologous gene family analysis of over 58,000 translated transcripts, implemented through Tribe-MCL, estimated the shared and unique gene space among the four species. This revealed 2025 conserved gene families, of which 408 were aligned to estimate levels of divergence and reveal patterns of selection. Specific candidate genes previously associated with drought tolerance and white pine blister rust resistance in conifers were investigated.