A consensus map of chromosome 6R in rye (Secale cereale L.)

Four F₂ mapping populations derived from crosses between rye inbred lines DS2×RXL10, 541×Ot1-3, S120×S76 and 544×Ot0-20 were used to develop a consensus map of chromosome 6R. Thirteen marker loci that were polymorphic in more than one mapping population constituted the basis for the alignment of the four maps using the JoinMap v. 3.0 software package. The consensus map consists of 104 molecular marker loci including RFLPs, RAPDs, AFLPs, SSRs, ISSRs, SCARs, STSs and isozymes. The average distance between the marker loci is 1.3 cM, and the total map length is 135.5 cM. This consensus map may be used as a source of molecular markers for the rapid development of new maps of chromosome 6R in any mapping population.     

A genetic linkage map of rye (Secale cereale L.)

 A genetic linkage map of rye composed of 91 loci (88 RFLP, two morphological and one isozyme markers) has been developed using two reciprocal crosses. The RFLP loci covering all seven chromosomes were detected by a selection of rye, wheat, barley and oat cDNA and genomic DNA probes. The level of polymorphism was dependent on the source of the clones, with a ranking of rye>wheat>barley>oat. Distorted segregations were detected in linkage groups of chromosomes 1R, 4R, 5R and 7R. When the recombination of the two reciprocal crosses was compared, no systematic increase or decrease in one or the other direction was observed suggesting that a combination of populations of reciprocal crosses is possible. Received: 5 August 1997/Accepted: 2 September 1997     

Genetic linkage map of rye (Secale cereale L.)

Communicated by G. Wenzel     

RFLP-based genetic map of rye (Secale cereale L.) chromosome 1R

Summary A map of chromosome 1R of rye was constructed using 16 molecular and biochemical loci. From long arm to short arm, known-function loci were placed in the order: XAdh — XLee — Glu-R1[Sec-3] — XPpdk-1R — XEm-1R-1 — XEm-1R-2 — Centromere — XNor-R1 —Gpi-R1 — XGli-R1 [Sec-1a] along with six anonymous genomic and cDNA clones from wheat. The map, which spans 106 cM with 12 loci clustered in a 15-cM region around the centromere, shows reasonably good agreement with previously published maps for the centromeric region, whereas the XNor-R1 — Gpi-R1 region gives a much larger distance than previously reported.     

A consensus linkage map of rye (Secale cereale L.) including 374 RFLPs, 24 isozymes and 15 gene loci

 Consensus linkage maps were constructed for all seven rye chromosomes using 12 basic RFLP maps. The maps presented contain a total of 413 markers. The number of markers per chromosome varies from 41 (chromosome 3R) to 83 (chromosome 1R). In addition to 374 RFLP and 24 isozyme markers 15 gene loci were incorporated, determining the traits reduced plant height, self fertility, male sterility restoration, vernalization response, resistance against powdery mildew, chlorophyll deficiency, hairy leaf sheath, hairy peduncle, waxy endosperm, waxless plant and absence of ligules. The maps presented allow the selection of markers for the fine mapping of certain regions of the rye genome. In terms of the known chromosomal rearrangements within the Triticeae its utilization can also be extended for mapping in wheat and barley. Received: 13 February 1998 / Accepted: 26 May 1998     

Embryogenesis and germination in rye (Secale cereale L.)

Summary Fine structural investigations of non-viable rye grains indicate recognisible abnormalities in the plasmalemma and mitochondrial membranes of the unimbibed embryo. Once such grains are wetted there is rapid and progressive disorganisation of the tissue. Biochemical studies show a reduced uptake of water, lack of respiratory activity and a failure in nucleic acid and protein synthesis. Whereas total DNA, RNA and protein levels are unchanged on loss of viability, the integrity of DNA and RNA is impaired and ribosomal RNA and soluble protein levels are reduced.     

Embryogenesis and germination in rye (Secale cereale L.)

Summary When rye embryos imbibe water they rapidly return to a condition of biochemical and structural complexity. Three stages of imbibition can be recognised: Phase I a short period (10 min) of physical wetting; Phase II a longer period (1 h) when little further imbibition occurs, followed by Phase III a continuous phase of active water uptake. The latter coincides with an increase in respiration rate and an increase both in the number of mitochondria and of cristae within them. Changes in fine structure become evident in all organelles in Phase III, after 2 h of imbibition. In the unimbibed embryo endoplasmic reticulum is present only as short crescents associated with electron lucent bodies, but in Phase III the endoplasmic reticulum proliferates to form many surrounding cirlets. After 6 h these circlets become fewer and instead the endoplasmic reticulum is seen in close association with the nuclear membrane. Concurrently incorporation of radioactive uridine and thymidine is first detectible. This suggests that the large increase in protein synthesis occurs on new ribosomes present on the reticulum associated with the nuclear membrane. For the first 6 h protein synthesis must occur either on polysomes within the dense packing of ribosomes or on these circlets of endoplasmic reticulum associated with electron lucent bodies.     

Molecular linkage mapping in rye (Secale cereale L.)

A rye linkage map containing clones from rye, wheat, barley, oat and rice genomic and cDNA libraries, known-function genes and microsatellite markers, was created using an F₂ population consisting of 110 F₂-derived F₃ families. Both co-dominant and dominant markers were added to the map. Of all probes screened, 30.8% were polymorphic, and of those polymorphic 79.3% were mapped. The current map contains 184 markers present in all seven linkage groups covering only 727.3 cM. This places a marker about every 3.96 cM on average throughout the map; however, large gaps are still present. The map contains 60 markers that have been integrated from previous rye maps. Surprisingly, no markers were placed between the centromere and C1–1RS in the short arm of 1R. The short arm of chromosome 4 also lacked an adequate number of polymorphic markers. The population showed a remarkable degree of segregation distortion (72.8%). In addition, the genetic distance observed in rye was found to be very different among the maps created by different mapping populations. Received: 10 January 2000 / Accepted: 26 May 2000     

Embryogenesis and germination in rye (Secale cereale L.)

Summary Cells of the young embryo contain highly differentiated organelles. During maturation and dehydration, complexity is reduced, the many layers of endoplasmic reticulum associated with electron lucent bodies become reduced to a few residual crescents, lipid droplets distributed in the cytoplasm migrate to and become closely appressed to the plasmalemma, mitochondrial cristae are reduced in number and dictyosomes are compacted.     

B-chromosomes in inbred lines of rye (Secale cereale L.)

B chromosomes from an experimental population of the Japanese JNK strain of rye, isogenic for its Bs, have been backcrossed into twelve different inbred lines. The experiment is a way of studying the effects of the Bs against a range of different homozygous A chromosome backgrounds. This publication deals with pairing effects of both the As and the Bs, and their interactions, and with pollen mitosis. At meiosis there is a genotypic component to B effects, and they do not appear to act solely through a physical disturbance within the nucleus. In pollen the Bs are always present in more than 50% of the grains regardless of their pairing behaviour during meiosis; this result fits with a parasitic model of the activity of rye Bs.     

Agrobacterium tumefaciens-mediated genetic transformation of rye (Secale cereale L.)

A system for the genetic transformation of rye by co-cultivation with Agrobacterium tumefaciens is described. A total of 45 independent transgenic plants were regenerated with a transformation efficiency of 1 to % of the inoculated explants. The co-cultivation of Agrobacterium-strain AGL0, harboring plasmid pJFnptII and rye im-mature embryos in liquid medium allowed a high throughput and facilitated washing of the cultures to avoid Agrobacterium overgrowth. Transgenic plants were phenotypically normal and fully fertile, which might be aconsequence of the short time in tissue culture. The selection with paromomycin exclusively during the regen-eration allowed the efficient recovery of transgenic events without interfering with somatic embryogenesis. Southern blot analysis confirmed the independent nature of the analyzed plants and indicated single copy inserts in more than 50% of them. Segregation analysis confirmed single locus integration and stable transgene expression in most of the lines, while one line with multiple locus integration was also observed. The analysis of T-DNA:: plant DNA boundary sequences revealed examples of exclusion of vector sequences, deletion of a few bases of the T-DNA or insertion of up to 29 bases of the vector backbone. This stresses the importance of detailed analysis of the inserted transgenes in order to identify events with the desired integration profile.     

B-chromosomes in inbred lines of rye (Secale cereale L.)

B-chromosomes from an experimental population of the Japanese JNK strain of rye, isogenic for its Bs, have been backcrossed into twelve different inbred lines. The experiment provides a way to study the effects of the Bs against a range of homozygous A-chromosome backgrounds. This publication deals with vigour and fertility: it shows that the rye Bs fit a parasitic model, and that they interact in their effects with the A-chromosome background genotype.     

Studies on enzymatic polymorphism in rye (Secale cereale L.) cultivars

An electrophoretic study of seed peroxidases of eight rye (Secale cereale L.) cultivars was carried out in order to estimate the polymorphism and the variability degree of this highly variable enzymatic system. Analysis of the Cathodal Peroxidase System (CPX) were performed using polyacrylamide gel electrophoresis. Thirteen loci were recorded in embryo plus scutellum and endosperm. The percentage of polymorphic loci ranged from 53.84% to 92.30% when the 99% polymorphism criterion was used; these high values were possibly due to the studied system. The average heterozygosity values ranged from 0.179 to 0.378; these values fell into the expected for open-pollinated species. Genetic distance values fitted well with those expected for conspocific plants with a little of diversification.     

Linkage relationships of esterase loci in rye (Secale cereale L.)

Summary Genetic analysis of esterase polymorphism in rye inbred lines with isoelectric focusing in polyacrylamide flat gels yielded evidence for the existence of at least ten esterase loci, Est 1–Est 10. The loci can be attributed to four different linkage groups (Est 1/ Est 2/Est 3/Est 5/Est 6/Est 7), (Est 4), (Est 8/Est 9), and (Est 10). Loci Est 5/Est 6/Est 7 and Est 8/Est 9, respectively, are tightly linked with a maximum recombination frequency of 0.2% and can therefore be regarded as compound loci which possibly originated in tandem duplications.     

Genetic control of aluminium tolerance in rye (Secale cereale L.)

 Aluminium (Al) tolerance in roots of two cultivars (“Ailés” and “JNK”) and two inbred lines (“Riodeva” and “Pool”) of rye was studied using intact roots immersed in a nutrient solution at a controlled pH and temperature. Both the cultivars and the inbred lines analysed showed high Al tolerance, this character being under multigenic control. The inbred line “Riodeva” was sensitive (non-telerant) at a concentration of 150 μM, whereas the “Ailes” cultivar showed the highest level of Al tolerance at this concentration. The segregation of aluminium-tolerance genes and several isozyme loci in different F₁s, F₂s and backcrosses between plants of “Ailés” and “Riodeva” were also studied. The segregation ratios obtained for aluminium tolerance in the F₂s analysed were 3 : 1 and 15 : 1 (tolerant : non-tolerant) while in backcrosses they were 1 : 1 and 3 : 1. These results indicated that Al tolerance is controlled by, at least, two major dominant and independent loci in rye (Alt1 and Alt3). Linkage analyses carried out between Al-tolerance genes and several isozyme loci revealed that the Alt1 locus was linked to the aconitase-1 (Aco1), nicotinamide adenine dinucleotide dehydrogenase-2 (Ndh2), esterase-6 (Est6) and esterase-8 (Est8) loci, located on chromosome arm 6RL. The order obtained was Alt1-Aco1-Ndh2-Est6-Est8. The Alt3 locus was not linked to the Lap1, Aco1 and Ndh2 loci, located on chromosome arms, 6RS, 6RL and 6RL respectively. Therefore, the Alt3 locus is probably on a different chromosome. Received: 18 March 1997 / Accepted: 21 March 1997     

Molecular analysis of mitochondrial DNA from rye (Secale cereale L.)

Summary Molecular characterization of mitochondrial (mt) DNA of rye (Secale cereale L.), free of significant amounts of contaminating chloroplast (cp) DNA, was initiated using the open-pollinated cultivar Halo as a source of mtDNA. Based on the compilation of data from restriction patterns, the molecular size of the mtDNA was estimated to be 410 Kb and its buoyant density was determined as 1.705 g/ml. Southern hybridization, using labelled cp genes (P700 and ribulosebiphosphate-carboxylase large subunit), indicated the presence of cpDNA-homologous regions on putative mtDNA fragments. Mt DNAs of inbred lines with fertile and cytoplasmic male sterile (CMS) Pampa cytoplasm were also analysed. Whereas the restriction patterns of mtDNAs of Halors and the fertile line turned out to be identical, Pampa mtDNA showed a unique restriction pattern, indicating (as in most other CMS systems) the involvement of mtDNA rearrangements in the expression of male sterility in rye. All 3 mtDNAs investigated contain regions homologous to the plasmid S1 of the CMS-S cytoplasm of Maize (Zea mays), as indicated by hybridization experiments. In Pampa cytoplasm the S-homologous sequence is located within a rearranged region of mtDNA.     

Linkage mapping of mutant loci in rye (Secale cereale L.)

Eight mutant loci determining the traits waxy plant (w and wa1), brown culm (cb), multiple pistils (mp), weak plant with reduced plant height (np), monoculm growth habit (mc), compactum growth habit (ct3) and anthocyaninless (an) were mapped on rye chromosomes 4R (w, np), 6R (cb, mc) and 7R (mp, wa1, ct3, an). For five mutants (w, wa1, cb, mp, np) molecular and biochemical markers were applied, whereas for mc, ct3 and an a classical linkage analysis was performed. Furthermore, it could be demonstrated that homoeologous relationships exist between most of the mapped rye loci and comparable mutants in wheat and barley. It was confirmed not only that genes controlling fundamental aspects of plant biology are highly conserved across the Triticeae species but so also were many mutant loci. Received: 19 June 2000 / Accepted: 18 October 2000     

Development and application of EST-based markers specific for chromosome arms of rye (Secale cereale L.)

To develop a set of molecular markers specific for the chromosome arms of rye, a total of 1,098 and 93 primer pairs derived from the expressed sequence tag (EST) sequences distributed on all 21 wheat chromosomes and 7 rye chromosomes, respectively, were initially screened on common wheat 'Chinese Spring' and rye cultivar 'Imperial'. Four hundred and fourteen EST-based markers were specific for the rye genome. Seven disomic chromosome addition lines, 10 telosomic addition lines and 1 translocation line of 'Chinese Spring-Imperial' were confirmed by genomic in situ hybridization and fluorescencein situ hybridization, and used to screen the rye-specific markers. Thirty-one of the 414 markers produced stable specific amplicons in 'Imperial', as well as individual addition lines and were assigned to 13 chromosome arms of rye except for 6RS. Six rye cultivars, wheat cultivar 'Xiaoyan 6' and accessions of 4 wheat relatives were then used to test the specificity of the 31 EST-based markers. To confirm the specificity, 4 wheat-rye derivatives of 'Xiaoyan 6 × German White', with chromosomes 1RS, 2R and 4R, were amplified by some of the EST-based markers. The results indicated that they can effectively be used to detect corresponding rye chromosomes or chromosome arms introgressed into a wheat background, and hence to accelerate the utilization of rye genes in wheat breeding.