No abstract

A BAC library of hexaploid wheat was constructed using the spring wheat cultivar Triticum aestivum L. 'Glenlea'. Fresh shoot tissue from 7- to 10-day-old seedlings was used to obtain HMW DNA. The library was constructed using the HindIII site of pIndigoBAC-5 and the BamHI site of pIndigoBAC-5 and pECBAC1. A total of 12 ligations were used to construct the entire library, which contains over 650 000 clones. Ninety-six percent of the clones had inserts. The insert size ranged from 5 to 189 kb with an average of 79 kb. The entire library was gridded onto 24 high-density filters using a 5 x 5 array. A subset of these membranes was hybridized with two intergenic chloroplast probes and the percentage of clones containing chloroplast DNA (cpDNA) was calculated to be 2.2%. The genome coverage was estimated to be 3.1 x haploid genome equivalents, giving a 95.3% probability of identifying a clone corresponding to any wheat DNA sequence. BAC pools were constructed and screened using markers targeting the Glu-B1 locus (1BL), the hardness loci (5AS, 5BS, 5DS), the leaf rust resistance locus Lr1 (5DL), and the major fusarium head blight QTL locus located on 3BS. These markers were either locus-specific amplicons or microsatellites. A total of 49 BAC clones were identified for 14 markers giving an average of 3.5 clones/marker, thereby corroborating the estimated 3.1x genome coverage. An example using the gene encoding the HMW glutenin Bx7 is illustrated.

Leaf rust (caused by Puccinia triticina Eriks.) is a worldwide and yield‐limiting disease of common wheat (Triticum aestivum L.). The gene Lr32, which confers seedling resistance to leaf rust, was previously transferred from Aegilops tauschii Coss. (RL5497‐1) to chromosome 3DS of common wheat. Without exception, Lr32 conferred resistance to 616 diverse isolates of leaf rust sampled from Canadian wheat fields over the past decade (2000 to 2009). Marker alleles specific to chromosome 3DS of RL5497 were found in Lr32 backcross lines of both Katepwa (BW196R) and Thatcher (RL6086). These introgressions from A. tauschii had no adverse effects on either agronomic performance or end use quality. A map based on 338 doubled haploid lines (DHL) from the cross of BW196R × Thatcher showed that Lr32 was slightly distal (0.6 cM) to two simple sequence repeat (SSR) loci wmc43 and barc135, which cosegregated on chromosome 3DS. A survey of the fragments amplified in wheats of diverse origin showed that both wmc43 and barc135 were cross‐applicable as markers for stacking Lr32 with other broadly effective leaf rust resistance genes.

The soybean cyst nematode (SCN), Heterodera glycines Ichinohe, is the foremost pest of soybean (Glycine max L. Merr.). The rhg1 allele on linkage group (LG) G and the Rhg4 allele on LG A2 are important in conditioning resistance. Markers closely linked to the Rhg4 locus were used previously to screen a library of bacterial artificial chromosome (BAC) clones from susceptible 'Williams 82' and identified a single 150-kb BAC, Gm_ISb001_056_G02 (56G2). End-sequenced subclones positioned onto a restriction map provided landmarks for identifying the corresponding region from a BAC library from accession PI 437654 with broad resistance to SCN. Seventy-three PI 437654 BACs were assigned to contigs based upon HindIII restriction fragment profiles. Four contigs represented the PI 437654 counterpart of the 'Williams 82' BAC, with PCR assays connecting these contigs. Some of the markers on the PI 437654 contigs are separated by a greater physical distance than in the 'Williams 82' BAC and some primers amplify bands from BACs in the mid-portion of the connected PI 437654 BAC contigs that are not amplified from the 'Williams 82' BAC. These observations suggest that there is an insertion in the PI 437654 genome relative to the 'Williams 82' genome in the Rhg4 region.

The sweet potato Ipomoea batatas (L.) Lam. is classified in series Batatas (Choisy) in Convolvulaceae, with 12 other species and an interspecific true hybrid. The phylogenetic relationships of a sweetpotato cultivar and 13 accessions of Ipomoeas in the series Batatas were investigated using the nucleotide sequence variation of the nuclear-encoded β-amylase gene. First, flowers were examined to identify the species, and DNA flow cytometry used to determine their ploidy. The sweetpotato accession was confirmed as a hexaploid, I. tabascana a tetraploid, and all other species were diploids. A 1.1–1.3 kb fragment of the β-amylase gene spanning two exons separated by a long intron was PCR-amplified, cloned, and sequenced. Exon sequences were highly conserved, while the intron yielded large sequence differences. Intron analysis grouped species currently recognized as A and B genome types into separate clades. This grouping supported the prior classification of all the species, with one exception. The species I. tiliacea was previously classified as a B genome species, but this DNA study classifies it as an A genome species. From the intron alignment, sequences specific to both A and B genome species were identified. Exon sequences indicated that I. ramosissima and I. umbraticola were quite different from other A genome species. Placement of I. littoralis was questionable: its introns were similar to other B genome species, but exons were quite different. Exon evolution indicated the B genome species evolved faster than A genome species. Both intron and exon results indicated the B genome species most closely related to sweetpotato (I. batatas) were I. trifida and I. tabascana.