A 34K SNP genotyping array for Populus trichocarpa: design, application to the study of natural populations and transferability to other Populus species.

Genetic mapping of quantitative traits requires genotypic data for large numbers of markers in many individuals. For such studies, the use of large single nucleotide polymorphism (SNP) genotyping arrays still offers the most cost-effective solution. Herein we report on the design and performance of a SNP genotyping array for Populus trichocarpa (black cottonwood). This genotyping array was designed with SNPs pre-ascertained in 34 wild accessions covering most of the species latitudinal range. We adopted a candidate gene approach to the array design that resulted in the selection of 34 131 SNPs, the majority of which are located in, or within 2 kb of, 3543 candidate genes. A subset of the SNPs on the array (539) was selected based on patterns of variation among the SNP discovery accessions. We show that more than 95% of the loci produce high quality genotypes and that the genotyping error rate for these is likely below 2%. We demonstrate that even among small numbers of samples (n = 10) from local populations over 84% of loci are polymorphic. We also tested the applicability of the array to other species in the genus and found that the number of polymorphic loci decreases rapidly with genetic distance, with the largest numbers detected in other species in section Tacamahaca. Finally, we provide evidence for the utility of the array to address evolutionary questions such as intraspecific studies of genetic differentiation, species assignment and the detection of natural hybrids.

Retroviral mediated gene transduction alters integrin expression on vascular endothelial cells.

Genetically recombinant endothelial cells (rEC) may improve the patency of small diameter vascular grafts by preventing thrombosis or limiting neointimal hyperplasia. Previous work has shown that rEC have reduced adhesion to vascular bypass grafts in vivo. Poor adhesion may be due to altered adhesion (integrin) receptors. This study evaluated the expression of the alpha 5 beta 1 (fibronectin), alpha 2 beta 1 (collagen IV), and alpha v beta 3 (vitronectin) integrin subunits on rEC. Human umbilical vein EC or canine jugular vein EC were transduced with neoR, neoR and human tPA or hygromycin resistance genes using retroviral vectors. Naive EC and EC exposed to empty viral particles (mEC) were controls. Naive EC, mEC, and all rEC's were evaluated for alpha and beta subunits for each integrin receptor studied using immunoblotting. Blotting for alpha 2, alpha 5, and alpha v exhibited expression of the alpha integrin subunits in all cells. The beta 1 and beta 3 subunits were present in mEC and nEC but were absent or truncated in all rEC. The decreased adhesion of rEC's to synthetic vascular grafts may be accounted for by their altered beta 1 and beta 3 integrin subunit expression. The beta subunit is critical for organization of the cytoskeleton and cellular signal transduction. Diminished beta subunit expression in rEC is neither vector specific nor related to retroviral exposure alone. Alteration of beta integrin expression may be to associated with the over-expression of phosphotransferase genes such as neoR or hygromycin B used as selectable markers in gene transfer protocols.