Alien Chromatin Research Articles

Molecular cytogenetic characterization of two high protein wheat-Thinopyrum intermedium partial amphiploids

Fluorescence and genomic in situ hybridization (FISH and GISH) were used to establish the cytogenetic constitution of two wheat × Thinopyrum intermedium partial amphiploids H95 and 55(1-57). Both partial amphiploids are high-protein lines having resistance to leaf rust, yellow rust and powdery mildew and have in total 56 chromosomes per cell. Repetitive DNA probes (pTa71, Afa family and pSc119.2) were used to identify the individual wheat chromosomes and to reveal the distribution of these probes within the alien chromosomes. FISH detected 6B tetrasomy in H95 and a null (1D)-tetrasomy (1B) in 55(1-57). GISH was carried out using biotin labeled Th. intermedium DNA and digoxigenin labeled Pseudoroegneria spicata DNA as probes, subsequently. GISH results revealed 44 wheat chromosomes and four Thinopyrum chromosome pairs, including three S and one J chromosome pairs in line H95. Line 55(1-57), contained 42 wheat chromosomes and six Th. intermedium pairs, including two S and one J(S) pairs. Additionally, two identical translocated chromosome pairs with diminished affinity to the alien chromatin were detected in both amphiploids. Another two translocations were found in 55(1-57), with satellite sections from the Thinopyrum J genome.

Construction of Identification System for Alien Chromatin in Crop Plant

In the previous decades, the expansion of genomic technologies has begun to revolutionize our ability to identify and measure genetic variation. The correct identification of alien chromatin involved in these gene transfers is crucial in a range of molecular and cytogenetic investigations. This paper presents the contribution of the sequential technique of C-banding, GISH and SSR marker on identification and detection of alien chromosomes, discusses the progress on identification of genetic variation, molecular and cytogenetic identification technologies, and prospects the significant construction of identification system for alien chromatin in crop plant.

The application of DNA marker and doubled-haploid technology for stacking multiple stem rust resistance genes in wheat

In wheat, the use of gene “pyramids” or “stacks” of major genes that confer resistance to all local strains of the fungal stem rust pathogen Puccinia graminis f. sp. tritici (Pgt) can increase durability of resistance where wheat cultivars with the single gene components are not widely deployed. Stacking two or more resistance genes becomes a breeding challenge, particularly when pathogen races that discriminate the genes are not available. The use of DNA markers and doubled-haploid technology provides a route for producing lines homozygous for multiple resistance genes. We have applied this approach to produce gene pyramids of two or more of the stem rust resistance genes Sr24 and new sources of SrR, Sr31 and Sr26 on reduced alien chromatin in the genetic backgrounds of Westonia and Pavon wheat. These genes, which are all derived from “alien” sources (SrR and Sr31 from rye, Sr24 and Sr26 from Agropyron elongatum) each provide resistance to all currently known pathotypes of Pgt in Australia, and SrR and Sr26 also provide resistance against all the variants of stem rust race Ug99 (TTKS group).

Sequence variations of PDHA1 gene in Triticeae species allow for identifying wheat-alien introgression lines

Abstract In order to develop a molecular marker for the detection of alien chromatin, an allele-specific primer targeting pyruvate dehydrogenase (PDHA1) gene was used to analyze 12 taxa representing different basic genomes in Triticeae. Ampli....cation products with different sizes were generated among species. The sequence alignments indicated that the PDHA1 genes contained some deletions/insertions of Miniature Inverted-repeat Terminal Elements (MITE) and simple sequence repeats (SSRs), thus suggesting that the Triticeae genomes have been rapidly evolving during speciation. The genome-specific amplicons and chromosomal location of PDHA1 gene on Triticeae genomes can be used to trace the corresponding alien chromatins from Aegilops, Secale and Dasypyrum species in wheat background.

Derivation and Characterization of Recombinants of the Lr54/Yr37 Translocation in Common Wheat

The wild relatives are an important source of new genes for the genetic improvement of wheat. Leaf and stripe rust resistance genes Lr54 and Yr37 occur on an Aegilops kotschyi-derived chromosomal translocation that had apparently replaced wheat chromosome arm 2DL. The alien chromatin also includes the locus of a gene for reduced plant height (H), which appears to be different from Rht8 on chromosome arm 2DS. The introgressed genes were mapped relative to homoeologous wheat marker loci following the induction of chromosome pairing in translocation heterozygotes that lacked the Ph1 locus. Ten recombined Lr54/Yr37 translocation chromosomes were derived and characterized with micro- satellite, AFLP and SCAR markers. The data suggested that there was significant homoeology between the full-length translocated segment and the wheat 2DL chromosome arm. The recombined translocations apparently resulted from single crossovers during which the distal end of the long arm of the translocation chromosome was replaced with wheat chroma- tin. Recombinant (Lr54/Yr37-74) retained the least alien chromatin and both resistance genes, yet had lost the reduced plant height gene. A polymorphic AFLP fragment was converted into a dominant SCAR marker to detect rec. #74. In addition three wheat microsatellite loci that map to the introgressed region provide a useful recessive marker system to detect Lr54/Yr37. The shortened translocation could be useful in breeding and may be used for continued, closer mapping of the resistance genes.

Chromosome Engineering of Durum Wheat with Alien Chromatin of Diploid Wheatgrass

Durum wheat (Triticum turgidum L., 2n = 4x = 28; AABB genomes) is an important cereal widely used for human consumption. It is the choice wheat for preparing pasta products. Current durum cultivars have little resistance to Fusarium head blight (FHB), or scab, a ravaging fungal disease caused by the fungal pathogen Fusarium graminearum Schwabe. We showed earlier that diploid wheatgrass (Lophopyrum elongatum (Host) Á. Löve (2n = 2x = 14; EE genome) is an excellent source of FHB resistance, which could be transferred to commercial durum cultivars by hybridization coupled with manipulation of chromosome pairing. We obtained homoeologous pairing (i.e., pairing between wheat and alien chromosomes) in intergeneric hybrids because of the wheatgrass-induced suppression of the pairing-regulating gene Ph1 of wheat, leading to alien chromatin integration into the durum genome. Adopting this chromosome engineering technology, we have transferred diploid wheatgrass chromatin into the durum genome. We have produced fertile hybrid derivatives with varying degrees of FHB resistance. Some derivatives showed only 7.6% infection compared with 72.0% in the parental durum cultivar. Fluorescent genomic in situ hybridization (fl-GISH) analysis was used to confirm the integration of alien chromatin into the durum complement.

Allosyndetic recombinants of theAegilops peregrina-derivedLr59translocation in common wheat

With 2 figures and 2 tables Abstract The wild relatives constitute a valuable source of rust resistance genes that can be utilized in wheat breeding. However, translocation of desirable genes through chromosome engineering inevitably results in co-transfer of deleterious wild species chromatin. An attempt was made to replace such redundant alien chromatin on the Lr59 translocation through homoeologous chromosome pairing and crossing over in the absence of Ph1. Strong segregation distortion resulted in the recovery of an unexpectedly high frequency of resistant recombinants. Eight of these retained comparatively short, distal segments of foreign chromatin, including Lr59. The foreign chromatin that remained in the latter plants was characterized with the use of twelve anonymous AFLP loci, the data of which suggested reduced homoeology with 1AL that could have been the result of a sub-terminal, paracentric inversion. Crossing over within an inversion loop may have resulted in a low frequency of genetically unbalanced gametes. It will therefore be necessary to develop near-isogenic lines of the eight recombinants and to do field evaluations in order to identify those that retained the shortest, balanced translocations.

Research progress in BYDV resistance genes derived from wheat and its wild relatives

Barley yellow dwarf virus (BYDV) may cause a serious disease affecting wheat worldwide. True resistance to BYDV is not naturally found in wheat. BYDV resistance genes are found in more than 10 wild relative species belonging to the genera of Thinopyrum, Agropyron, Elymus, Leymus, Roegneria, and Psathyrostachy. Through wide crosses combining with cell culture, use of ph mutants, or irradiation, 3 BYDV resistance genes in Th. intermedium, including Bdv2, Bdv3 and Bdv4, were introgressed into common wheat background. Various wheat-Th. intermedium addition and substitution, translocation lines with BYDV-resistance were developed and characterized, such as 7D-7Ai#1 (bearing Bdv2), 7B-7Ai#1, 7D-7E (bearing Bdv3), and 2D-2Ai-2 (bearing Bdv4) translocations. Three wheat varieties with BYDV resistance from Th. intermedium were developed and released in Australia and China, respectively. In addition, wheat-Agropyron cristatum translocation lines, wheat-Ag. pulcherrimum addition and substitution lines, and a wheat-Leymus multicaulis addition line (line24) with different resistance genes were developed. Cytological analysis, morphological markers, biochemical markers, and molecular markers associated with the alien chromatin carrying BYDV resistance genes were identified and applied to determine the presence of alien, chromosomes or segments, size of alien chromosome segments, and compositions of the alien chromosomes. Furthermore, some resistance-related genes, such as RGA, P450, HSP70, protein kinases, centrin, and transducin, were identified, which expressed specifically in the resistance translocation lines with Bdv2. These studies lay the foundations for developing resistant wheat cultivars and unraveling the resistance mechanism against BYDV.

TaASY1 promotes homologous chromosome interactions and is affected by deletion of Ph1

During meiosis, chromosomes are sorted into homologous pairs as a preface to their intimate association via recombination and synapsis. However, little is known about the mechanism used to distinguish homologous chromosomes from other chromosomes present in the nucleus. Studies in wheat (Triticum aestivum) have shown that the Pairing homoeologous 1 (Ph1) locus is required to suppress interactions between genetically similar homoeologous chromosomes. Here we show that absence of Ph1 causes increased transcription of Asynapsis 1 (ASY1), a gene that encodes an axial-element-associated protein that is essential for synapsis and cross-over formation in Arabidopsis and rice. Localisation of ASY1 during meiosis is also affected by deletion of Ph1. In addition, transgenic wheat mutants with decreased activity of TaASY1 display reduced synapsis during prophase I and exhibit pairing between homoeologous chromosomes at metaphase I. These results suggest that ASY1 is required to promote interactions between homologous chromosomes in bread wheat, and that Ph1 has a gene regulatory role, which is consistent with its suggested genetic identity as a Cdk-like gene. Broader implications of this research suggest that we could use the Taasy1 mutants to assess their efficacy in alien chromatin introgression studies, as seen with the ph1b mutant.

Locating the alien chromatin segment in common wheat-Aegilops longissima mildew resistant transfers

Locating the alien chromatin segment in common wheat-Aegilops longissima mildew resistant transfers

Locating the alien chromatin segment in common wheat-Aegilops longissima mildew resistant transfers

By use of the Chinese Spring ph 1b mutant, recombination was induced between the 3S1 short arm telosome of Ae. longissima and its group-3 wheat homoeologues. Transfer was thus obtained of chromosomal segments bearing the alien Pm 13 mildew resistance gene into several common wheat lines. To identify the wheat chromosome involved in each transfer, these were subjected to monosomic and C-banding analyses. Probably due to preferential 3B-3S1 pairing, irrespective of the group-3 wheat chromosome present as monosome in the critical steps of the transfer work, in 2/3 of the cases 3B turned out to be the recipient chromosome, whereas 3D was the one involved in the remaining ones. Assessment of the residual pairing ability of the recombinant chromosomes in F1's between four 3B and three 3D transfer lines and their corresponding wheat ditelosomic as well as 3S1 substitution lines, indicated about coincident values (about 40 % pairing with DT3BS) in three of the 3B recombinants, a significantly different pattern in the fourth one (10 %) and a more continuous variation among the 3D ones. An overall prevailing terminal location of the wheat-alien chromatin exchange points is tentatively hypothesized.

Analysis of DNA methylation variation in wheat genetic background after alien chromatin introduction based on methylation-sensitive amplification polymorphism

During the process of alien germplasm introduced into wheat genome by chromosome engineering, extensive genetic variations of genome structure and gene expression in recipient could be induced. In this study, we performed GISH (genome in situ hybridization) and AFLP (amplified fragment length polymorphism) on wheat-rye chromosome translocation lines and their parents to detect the identity in genomic structure of different translocation lines. The results showed that the genome primary structure variations were not obviously detected in different translocation lines except the same 1RS chromosome translocation. Methylation sensitive amplification polymorphism (MSAP) analyses on genomic DNA showed that the ratios of fully-methylated sites were significantly increased in translocation lines (CN12, 20.15%; CN17, 20.91%; CN18, 22.42%), but the ratios of hemimethylated sites were significantly lowered (CN12, 21.41%; CN17, 23.43%; CN18, 22.42%), whereas 16.37% were fully-methylated and 25.44% were hemimethylated in case of their wheat parent. Twenty-nine classes of methylation patterns were identified in a comparative assay of cytosine methylation patterns between wheat-rye translocation lines and their wheat parent, including 13 hypermethylation patterns (33.74%), 9 demethylation patterns (22.76%) and 7 uncertain patterns (4.07%). In further sequence analysis, the alterations of methylation pattern affected both repetitive DNA sequences, such as retrotransposons and tandem repetitive sequences, and low-copy DNA.

AFLP and SCAR Markers Linked to Tomato Spotted Wilt Virus Resistance in Tobacco

Tomato spotted wilt virus (TSWV) is a serious disease in tobacco (Nicotiana tabacum L.). The breeding line ‘Polalta’ contains a single dominant gene conferring resistance to TSWV that was introgressed from N. alata Link & Otto. The resistance is tightly associated with an abnormal plant type, however, and traditional backcrossing has been ineffective in producing normal plants with TSWV resistance. A potential strategy to overcome this problem is to use molecular markers to select against alien chromatin in backcross progeny. Amplified fragment length polymorphism (AFLP) technology and bulked segregant analysis were applied to identify markers linked to the resistance gene. The DNA bulks from susceptible and resistant doubled haploid lines derived from a cross between susceptible cultivar K326 and Polalta were analyzed to identify linked markers. An F2 population and the doubled haploids were used to construct a 2.5‐cM map of this locus containing 17 coupling phase and seven repulsion phase markers. By selecting for resistant plants with an improved plant type and marker profile in a BC3 population, four plants were identified that were missing 1 to 14 of the AFLP markers associated with the alien chromatin, demonstrating that rare recombination events can be identified. Four AFLP fragments were successfully converted to sequence characterized amplified region markers suitable for large‐scale screening. This approach may facilitate development of TSWV‐resistant tobacco cultivars.

Use of fluorescence in situ hybridization as a tool for introgression analysis and chromosome identification in coffee (Coffea arabica L.)

Fluorescence in situ hybridization (FISH) was used to study the presence of alien chromatin in interspecific hybrids and one introgressed line (S.288) derived from crosses between the cultivated species Coffea arabica and the diploid relatives C. canephora and C. liberica. In situ hybridization using genomic DNA from C. canephora and C. arabica as probes showed elevated cross hybridization along the hybrid genome, confirming the weak differentiation between parental genomes. According to our genomic in situ hybridization (GISH) data, the observed genomic resemblance between the modern C. canephora genome (C) and the C. canephora-derived subgenome of C. arabica (Ca) appears rather considerable. Poor discrimination between C and Ca chromosomes supports the idea of low structural modifications of both genomes since the C. arabica speciation, at least in the frequency and distribution of repetitive sequences. GISH was also used to identify alien chromatin segments on chromosome spreads of a C. liberica-introgressed line of C. arabica. Further, use of GISH together with BAC-FISH analysis gave us additional valuable information about the physical localization of the C. liberica fragments carrying the SH3 factor involved in resistance to the coffee leaf rust. Overall, our results illustrate that FISH analysis is a complementary tool for molecular cytogenetic studies in coffee, providing rapid localization of either specific chromosomes or alien chromatin in introgressed genotypes derived from diploid species displaying substantial genomic differentiation from C. arabica.

Manitoba Regional Meeting, 2006 The Canadian Phytopathological Society

Effects of mutations in a bacterial biocontrol agent,Pseudomonas chlororaphis PA23, on its root-colonization ability and production of volatile compounds. S.N.P. Athukorata, W.G.D. Fernando, T.R. de kievet, and K.Y. Rashid. Department of Microbiology, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (K.Y.R.) Cereal Research Centre, Agriculture and Agri-Food Canada, Modern Research Station, Unit 100-101, Route 100, Modern, MB R6M 1Y5, Canada. Effects of bacterial and jasmonic acid treatments on insect pests of canola. K. Bergen, W.G.D. Fernando, and N.J. Holliday.Department of Plant Science, University of Manitoba, Winnipeg, MB R3T 2N2, Canada. The role of the stringent response in Pseudomonas spp. DF41 biocontrol. J. Brucal, W.G.D. Fernando, and T.R. de kievit. Department of Microbiology; and (W.G.D.F.) Department of Plant Science, University of Manitoba, Winnipeg, MB R3T 2N2, Canada. Population structure of the blackleg pathogen Leptosphaeria maculansfrom North America.Y.Chen, B. Fristensky, and W.G.D. Fernando. Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada. New effective sources of resistance to crown rust [Puccinia coronata f.sp avenae] in Avena sterilis. J.Chong and G. Chen. Cereal Research Centre, Agriculture and Agri-Food Canada, 195 Dafoe Road, Winnipeg, MB R3T 2M9, Canada. Breeding for canola resistance against blackleg caused by PG3 and PG4 strains of Leptosphaeria amculans. M. Dusabenyagasani and W.G.D. Fernando.Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada. The role of pycnidiospores as primary inoculum, and assessment of disease severity on canola or rapeseed (Brassica napus) Plants infected at different growth stages. k. Ghanbarnia and W.G.D. Fernando. Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada. Prediction of deoxynivalenol in spring wheat based on cropping practices,Fusarium spores on a wheat head, and rainfall at the flowering stage. X.W. Guo, W.G.D. Fernando, H. Sapirstein, P. Bullock, and T. Nowicki.Department of Plant Science; (H.S.) Department of Food Science; (P.B.) Department of Soil Science, University of Manitoba, MB R3T 2N2, Canada; and (T.N.) Grain Research Laboratory, Canadian Grain Commission, 1404-303 Main Street, Winnipeg, MB R3C 3G8, Canada. Breaking the linkage between the Wsm1 gene for resistance to Wheat streak mosaic virus and the alien chromatin of its origin.S. Haber, M. Pradhan, and D. Somers.Cereal Research Centre, Agriculture and Agri-Food Canada; and (M.P.) Department of Botany, University of Manitoba MB R3T 2N2, Canada. The role of pyrolnitrin produced by Pseudomonas chlororaphis PA23 in the biological control of Sclerotinia sclerotiorum in canola. R. Habibian, T.R. de Kievet, and W.G.D. Fernando.Department of Plant Science; and (R.H., T.R.K.) Department of Microbiology, University of Manitoba MB R3T 2N2, Canada. Molecular and biochemical detection of Bacillus spp. producing fengycin and bacillomycin D, Antagonistic to fungal Pathogens of canola and wheat. R. Ramarathnam, S. Bo, Y. Chen, W.G.D. Fernand, and G. Xuewen.Department of Plant Science, 222 Agriculture Building, University of Manitoba, Winnipeg, MB R3T 2N2, Canada; and (S.B., G.X.) Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural university, No. 1 Weigang Road, Nanjing 210095, China.

Molecular identification of Pm12-carrying introgression lines in wheat using genomic and EST-SSR markers

Pm12, transferred from Aegilops speltoides (2n = 2x = 14, genome SS) to wheat, confers effective resistance to powdery mildew worldwide. By applying bulked segregant analysis in a BC3F2 segregating population consisting of 305 plants, 18 wheat genomic and EST-SSR markers linked to the resistance gene were identified. Pm12 was located in the 6SS portion of the T6BS-6SS.6SL translocation chromosome based on the physical bin positions of the genomic and EST-SSR markers in the Chinese Spring group six deletion stocks and their linkage relationship to the resistance gene. Twenty eight recombinants among 305 F2 plants indicated a low frequency of recombination between the alien chromosome segment and wheat chromosome 6B. Since recombination events occurred on both sides of Pm12, the materials generated provide opportunities for further reduction of alien chromatin by intercrossing selected individuals and using markers to select the required plants.

Widening the gene pool of cultivated lentils through introgression of alien chromatin from wild Lens subspecies

AbstractWild Lens species/subspecies are a potential source for increasing genetic diversity in cultivated lentil. Four intraspecific crosses were attempted between cultivated and wild lentils. Viable hybrids were produced between L. culinaris ssp. culinaris × L. culinaris ssp. orientalis and L. culinaris ssp. culinaris × L. culinaris ssp. odomensis. Normal meiosis and pollen fertility were observed in the first set of crosses, whereas chromosomal abnormalities and reduced pollen fertility were observed in the second set of crosses. These crosses were also studied for some quantitative traits. The range, mean and coefficient of variation were calculated in parents, F1, F2 and BC1 generations to determine the extent of variability generated in the cultivated lentil through introgression of genes from wild lentil. The cultivated lentil × L. culinaris ssp. orientalis crosses showed substantially higher variability for all the traits than crosses involving cultivated lentil ×L. culinaris ssp. odomensis. The results of the present study indicated that these wild subspecies can be exploited for breeding purposes and their variation can easily be utilized to widen the genetic base of the cultivated lentil.

New sources of rust resistance from alien species: meliorating linked defects and discovery

This paper presents a review of projects undertaken over the past 2 decades aimed at improving the yield and/or quality attributes of translocation lines carrying rust resistance genes from species related to wheat, so as to make these lines more suitable for use in breeding programs. Homeologous recombination between the alien chromosome segments and normal wheat chromosomes was induced in a ph1bph1b background. Lines with shortened alien chromatin were selected using dissociation patterns of molecular-based markers. A new line of bread wheat was developed containing a shortened chromosome 1RS segment carrying rust resistance gene SrR (Secale cereale L.), in which a defect affecting dough-quality appears to have been deleted. In addition, several advanced lines were produced with modified 6Ae#1L chromosome segments with Sr26 (Thinopyrum ponticum), 2S#1 chromosome segments with Sr32, and a previously unnamed gene, a 2S#2 chromosome segment with Sr39 (Triticum speltoides), 4G#1 chromosome segments with Sr37, and 2G#2 chromosome segments with Sr40 (T. timopheevii).

Retroelement Genome Painting: Cytological Visualization of Retroelement Expansions in the Genera Zea and Tripsacum

Divergence of abundant genomic elements among the Zea and Tripsacum genera was examined cytologically and a tool kit established for subsequent studies. The LTR regions from the CRM, Huck, Grande, Prem1, Prem2/Ji, Opie, Cinful-1, and Tekay retroelement families were used as FISH probes on mitotic chromosome spreads from a "trispecies" hybrid containing chromosomes from each of three species: Zea mays (2n = 20), Z. diploperennis (2n = 20), and Tripsacum dactyloides (2n = 36). Except for Tekay, which painted both Zea and Tripsacum chromosomes with nearly equal intensity, the retroelement probes hybridized strongly to the Zea chromosomes, allowing them to be distinguished from those of Tripsacum. Huck and Grande hybridized more intensely to maize than to Z. diploperennis chromosomes. Tripsacum genomic clones containing retroelement sequences were isolated that specifically paint Tripsacum chromosomes. The retroelement paints proved effective for distinguishing different genomes in interspecific hybrids and visualizing alien chromatin from T. dactyloides introgressed into maize lines. Other FISH probes (180-bp knob, TR-1, 5S, NOR, Cent4, CentC, rp1, rp3, and alpha-ZeinA) could be simultaneously visualized with the retroelement probes, emphasizing the value of the retroelement probes for cytogenetic studies of Zea and Tripsacum.

Molecular and Genetic Characterization of Nicotiana glutinosa L. Chromosome Segments in Tobacco mosaic virus‐Resistant Tobacco Accessions

Tobacco mosaic virus (TMV)‐resistant flue‐cured tobacco (Nicotiana tabacum L.) cultivars have been developed using the N gene derived from N. glutinosa L. Their adoption has been low, however, because of unfavorable linkage drag effects. Strategies to overcome this problem might include pursuit of alternative introgression events and/or use of molecular markers for selection against deleterious alien chromatin. Previous workers demonstrated the presence of a TMV‐resistance mechanism on more than one chromosome of the tobacco genome. The objectives of this research were to determine the relative genomic positions of TMV resistance loci in a set of 12 TMV‐resistant tobacco accessions and to use amplified fragment length polymorphism (AFLP) markers for characterization of this material with respect to linked alien chromatin. Five accessions were found to carry a TMV resistance gene on chromosome H. Seven accessions were found to carry a resistance factor on an alternative chromosome. Polymerase chain reaction results indicated that the N gene from N. glutinosa is responsible for resistance in all 12 accessions. A set of 168 AFLP markers specific to the N. glutinosa donor chromosome was identified and used to reveal variability among the 12 accessions for the relative amounts of N. glutinosa chromatin linked to the N gene. The relative propensity for crossing over within the alien segment when in different genomic positions was evaluated in BC1F1 families derived from three different accessions. Lines possessing the N gene on chromosome H may be of greater practical value because of relatively smaller introgressed alien segments and increased potential for obtaining crossover events within the segments.