Geographic differentiation of domesticated einkorn wheat and possible Neolithic migration routes.

To analyse the spread of domesticated einkorn into Europe, 136 landraces, 9 wild einkorns and 3 Triticum urartu were fingerprinted by the diversity array technology sequence (DArT-seq) marker technology. The obtained 3455 single-nucleotide polymorphism (SNP) markers confirmed earlier results about the separation of wild and domesticated einkorn from T. urartu and about the pinpointing of the domesticated forms to the Karacadag Mountains (Turkey). Further analyses identified two major domesticated landrace einkorn groups, one relating to the Prealpine region and the other to the Maghreb/Iberian region. The previously published four geographical provenance groups were mostly identified in our results. The earlier reported unique position of the Maghreb/Iberia einkorns cannot be confirmed, as the three landrace clusters we identified with STRUCTURE also occur in the remaining einkorn, although at different frequencies. The results are discussed with respect to the spreading of domesticated einkorn into Western Europe and two possible Neolithic migration routes are indicated.

An AFLP-based procedure for the efficient mapping of mutations and DNA probes in barley.

A strategy based upon AFLP markers for high-efficiency mapping of morphological mutations and DNA probes to linkage groups in barley is presented. First, 511 AFLP markers were placed on the linkage map derived from the cross Proctor x Nudinka. Second, loci controlling phenotypic traits were assigned to linkage groups by AFLP analysis, using F2 populations consisting of 30-50 mutant plants derived from crosses of the type "mutant x Proctor" and "mutant x Nudinka." To map DNA probes, 67 different wild-type barley lines were selected to generate F2 populations by crossing with Proctor and Nudinka. F2 plants that were polymorphic for a given RFLP fragment were classified into genotypic classes. Linkage of the RFLP polymorphism to 1 of the 511 AFLP loci was indicated by cosegregation. The use of the strategy is exemplified by the mapping of the mutation branched-5 to chromosome 2 and of the DNA probes Bkn2 and BM-7 to chromosomes 5 and 1, respectively. Map expansion and marker order in map regions with dense clustering of markers represented a particular problem. A discussion considering the effect of noncanonical recombinant products on these two parameters is provided.

A comparison of AFLPs and microsatellites to identify the population structure of brown trout (Salmo trutta L.) populations from Hardangervidda, Norway.

A total of 495 fish from 11 Hardangervidda lakes were genotyped in order to compare amplified fragment length polymorphisms (AFLP) and microsatellites in terms of their capacity to infer population genetic structure. The 11 microsatellites used in this study gave a greater polymorphism information content and greater gene diversity, with an average of 14.8 alleles per locus, than the six AFLP primer combinations used. However, the AFLPs resulted in 178 polymorphic loci and a 3.1 times larger marker index (effective multiplex ratio multiplied with the gene diversity). Comparable population structuring, for example in terms of distinguishing fish from the different river systems, was obtained with both marker systems. An AFLP and microsatellite multilocus Bayesian assignment test with the structure program divided the fish into six groups largely concurrent with main branches on a population neighbour-joining tree. Yet, the admixture status of individuals is mostly contradictory in the AFLP and the microsatellite analyses. The results are discussed concerning migration between lake populations.

C-banding pattern and powdery mildew resistance of Triticum ovatum and four T. aestivum-T. ovatum chromosome addition lines.

C-banding patterns of T. ovatum (Ae. ovata) and four T. aestivum cv 'Poros'-T. ovatum chromosome addition lines are presented, and the added chromosomes of T. ovatum have been identified. Furthermore, nucleolar activity and powdery mildew resistance were analyzed in the 'Poros'-ovatum addition lines and compared to that of T. ovatum and T. aestivum cv 'Poros'. The addition lines II, III and IV and 'Poros' were highly susceptible to powdery mildew isolates nos. 8 and 9, whereas the addition lines VI1 and VI2 showed high resistance. Even for an Ml-k virulent isolate, these two lines were highly resistant. By combining the cytological results and those of the powdery mildew analysis, the added chromosomes of T. ovatum can be excluded from responsibility for the high powdery mildew resistance of the addition lines VI1 and VI2. The same is true for a modified chromosome 6B, which is present in the 'Poros'-ovataum addition lines II, III and VI. The high variation in C-banding pattern observed in the A-, B- and D-genome complement of the addition lines is believed to be the result of crossing different lines of T. aestivum instead of 'Poros' alone. Thus, we cannot trace the powdery mildew resistance back to a specific chromosome.

Mapping of digested and undigested random amplified microsatellite polymorphisms in barley.

The broad use of microsatellites as a tool for constructing linkage maps in plants has been limited by the need for sequence data to detect the underlying simple sequence repeats. Therefore, random amplified microsatellite polymorphisms (RAMPs) were studied as an alternative approach for barely mapping. Labelled (GA)n simple sequence repeat primers were combined with RAPD primers of different length and sequence to generate RAMPs. To get additional polymorphisms (called dRAMPs), the obtained products were also analysed after digestion with MseI. There were 0-11 polymorphisms found per primer combination. Sixty RAMPs/dRAMPs identifying 40 new loci were mapped onto a barley RFLP map. The new DNA markers are found on all chromosomes and they increased the length of the barely map by 174 cM to a total of 1270 cM. Interestingly, the RAMPs/dRAMPs caused stretching effects in genome areas where stretching was also observed for AFLPs.

Barley microsatellites: allele variation and mapping.

Microsatellites have developed into a powerful tool for mapping mammalian genomes and first reports about their use in plants have been published. A database search of 228 barley sequences from GenBank and EMBL was made to determine which simple sequence repeat (SSR) motif prevails in barley. Nearly all types of SSRs were found. The (A)n and (T)n SSRs occurred more often than (C)n and (G)n for n > or = 10. Among the dinucleotide repeats, the (CG)n SSRs occurred least often. Trinucleotide repeats did not occur with n > 7 and there is no correlation between the GC content in the trinucleotide motifs and the number of observed SSRs. Analysing 15 different microsatellites with 11 barleys yielded 2.1 alleles per microsatellite. Sequencing 25 putative microsatellites showed that the resolution capacity of high-quality agarose gels was sufficient to determine differences of only three base pairs. Five microsatellites were mapped on three different chromosomes of a barley RFLP map.

Inheritance of RAPDs in F1 hybrids of corn.

Random amplified polymorphic DNA (RAPD) markers were analyzed in materials from a partial diallel, including 16 corn F1 hybrids (with five reciprocals) and their five parental inbreds. Using 21 primers, we scored a total of 140 different fragments for their presence/absence and intensity variation, where appropriate. When all 21 genotypes were taken into consideration, 20.7% of these fragments were nonpolymorphic, 37.1% were unambiguously polymorphic, and 42.1% were quantitatively polymorphic. Unambiguous polymorphisms were distinguished by the simple presence or absence of a specific fragment in the inbred genotypes, whereas quantitative polymorphisms exhibited a variation in the intensity of a fragment. Of the F1 patterns, 95.2% of the unambiguously polymorphic situations could be interpreted genetically by assuming complete dominance of the presence of the parental fragment, while 3.2% of the F1 patterns exhibited a fragment intensity that was intermediate between the two parental patterns (partial dominance). For quantitative polymorphisms, values of 88.1% for complete dominance and 5.0% for partial dominance were obtained. The results suggest that specific types of errors can be detected in RAPD analysis, that uniparental inheritance is not common, and that RAPD analysis might be more prudently used for some applications than for others.

Cytological characterization, powdery mildew resistance and storage protein composition of tetraploid and hexaploid 1BL/1RS wheat-rye translocation lines.

Progenies of a tetraploid 1BL/1RS wheat-rye translocation line, CV 256, selected from the cross 'Cando' x 'Veery', were analyzed by means of Giemsa C-banding. CV 256 is cytologically stable for the presence of the 1BL/1RS translocation but still segregating for A- and B-genome chromosomes of 'Cando' and 'Veery'. In CV 256, nucleolar activity of the 1RS NOR locus is suppressed, as judged by the absence of a secondary constriction in that rye segment and the capability of organizing nucleoli. PAGE analysis of prolamins confirmed the presence of two 1RS secalins in all single seeds analyzed. SDS-PAGE analysis of reduced glutenins of single seeds indicated that some seeds contained the 'Cando' Glu-B1 locus (subunits 6+8), some contained the 'Veery' Glu-B1 locus (subunits 7+9) while others contained all four subunits, indicating that the material was heterozygous. Pm8 resistance is expressed in the tetraploid 1BL/1RS translocation line based on the reactions of six well-defined powdery mildew isolates. However, Pm8 resistance is not expressed in the hexaploid wheat cultivars 'Olymp', 'Heinrich' and 'Florida', which also contain the 1BL/1RS translocation. Obviously, the existence of the 1BL/1RS translocation is not a proof for the expression of the associated genes. PAGE results did not show a clear linkage between powdery mildew resistance and the presence of 1RS secalins.

The incorporation and characterization of powdery mildew resistance from Aegilops longissima in common wheat (T. aestivum L.).

In the progeny of a hybrid between monotelosomic line 3B of 'Chinese Spring' wheat and 'Chinese Spring' - Aegilops longissima ditelosomic addition line 'G' a cytologically stable strain was selected consisting of 20 wheat chromosome pairs, one pair of telosomic chromosome 3BL and one pair of telosomic longissima chromosome 'G'. Inoculating 'Chinese Spring' - Aegilops longissima addition and substitution lines with ten different powdery mildew isolates, partial resistance was observed. The infection grade as well as the number of spores/cm(2) leaf area were significantly reduced.

Mapping maize microsatellites and polymerase chain reaction confirmation of the targeted repeats using a CT primer.

Microsatellites, also called simple sequence repeats (SSRs), have yielded an important class of DNA markers most notable for mapping mammalian genomes. To study the occurrence of microsatellites and their inheritance in maize, a search was made of 280 maize GenBank sequences. Six SSRs were chosen and unique flanking primers were designed for polymerase chain reaction (PCR) amplification. Eight different maize inbreds were studied with these six primer pairs and a mean of 3.5 polymorphic patterns occurred within the expected size range. For five of these putative microsatellites, the segregation in a maize restriction fragment length polymorphism mapping population was analyzed. Four of the microsatellites cosegregated with the Adh1, Gpc1, Pdk1, and Tpi genes from which the primer sequences were derived. The fifth primer pair (MZEGPA1) showed segregating polymorphisms, but the products were larger than expected. To verify the existence of the original SSRs in the segregating PCR products, a CT primer, containing a CT SSR and an arbitrary leader sequence, was used to reamplify these products. The four microsatellites that cosegregated with the original gene were reamplified as anticipated, whereas a suspicious 230-bp product obtained when using the MZEGPA1 primers could not be reamplified. Based on these results it is concluded that microsatellites can be a valuable tool for maize mapping.

A comparison of RAPD and isozyme analyses for determining the genetic relationships among Avena sterilis L. accessions.

Isozyme analysis is a valuable tool for determining genetic relationships among breeding lines and populations. The recently developed DNA technologies which can assay a greater proportion of the plant genome are providing a plentiful array of additional genomic markers. The objective of this research was to compare random amplified polymorphic DNA (RAPD) versus isozyme-based estimation of relationships among 24 accessions of a hexaploid wild oat, Avena sterilis L. The accessions were evaluated for variation in 23 enzyme systems and by 21 10-mer primers. A total of 77 polymorphic isozyme bands and 115 polymorphic RAPD bands were observed. Two matrices of genetic distances were estimated based on band presence/ absence. These matrices were subsequently utilized in cluster analysis and principal coordinate analysis. Both isozymes and RAPDs were proficient at distinguishing between the 24 accessions. The correspondence between the elements of both distance matrices was moderate (r=0.36**). Nevertheless, the overall representation of relationships among accessions by cluster analysis and ordination was in considerable agreement. The two techniques contrasted most notably in pair-by-pair comparisons of relationships. RAPD analysis resulted in a more definitive separation of clusters of accessions. The most significant impact of the DNA-based markers probably will be the more accurate determination of relationships between accessions that are too close to be accurately differentiated by isozymes.

Intrachromosomal mapping of seven biochemical loci in barley, Hordeum vulgare.

Seven biochemical loci, AmpA, Amy1, Amy2, Est-H5, Hor1, Hor2, and Wsp-H1, have been intrachromosomally mapped in the barley genome using a previously published RFLP-based genetic map. In all cases, the map locations confirmed prior chromosome assignments and agreed closely with the map positions of their homoeoloci in hexaploid wheat.

RFLP-based genetic relationships of Einkorn wheats.

To study the relationships between different species of the Einkorn group, 55 different accessions ofTriticum monococcum,T. boeoticum,T. urartu,T. sinskajae,T. thaoudar andT. aegilopoides were analyzed. Fifteen anonymous probes and four clones corresponding to storage protein genes were used for detecting restriction fragment length polymorphisms (RFLPs). The DNA was restricted with the restriction enzymesAluI,HaeIII,RsaI andTaqI. The 25 probe/enzyme combinations employed yielded a total of 488 polymorphic fragments. Statistical analyses were performed using Jaccard's coefficient of similarity and principal coordinate analysis. Different values of similarity within the three main taxa,monococcum,boeoticum andurartu, were obtained; the grouping at the species level was quite well reflected by the RFLP analysis done here. The coincidence between RFLP data and the subspecies classification of theT. monococcum group was only partial. OneT. urartu accession is clearly different from all of the other 54 accessions. The need for an RFLP based revision of the Einkorn taxonomy is evident.

Comment on "AFLP data and the origins of domesticated crops".

We review some concepts and methods of handling and using DNA fingerprinting in phylogenetic analyses related to crop domestication. Particular reference is made to AFLP markers and mode and place of einkorn, barley, and tetraploid wheat domestication in the Neolithic by human communities in the Fertile Crescent. The reconsideration of AFLP databases of domesticated and wild lines demonstrates that phylogenetic tree topologies, originally described for the three species, match closely the new results obtained by principle coordinate analyse.

Combined mapping of AFLP and RFLP markers in barley.

AFLP marker technology allows efficient DNA fingerprinting and the analysis of large numbers of polymorphic restriction fragments on polyacrylamide gels. Using the doubled haploids from the F1 of the cross Proctor x Nudinka, 118 AFLP markers were mapped onto a barley (Hordeum vulgare L.) RFLP map, also including five microsatellite and four protein marker loci. The AFLP markers mapped to all parts of the barley chromosomes and filled in the gaps on barley chromosomes 2L, 4L and 6 in which no RFLP loci had been mapped. Interestingly, the AFLP markers seldom interrupted RFLP clusters, but grouped next to them. The combined map covers 1873 cM, with a total of 282 markers. The merging of AFLP and RFLP markers increased the total map length; 402 cM were added to the map at the tips of chromosomes or in regions corresponding to earlier gaps. Another 375 cM resulted from mapping AFLP markers near to RFLP clusters or in between non-clustered RFLP markers.

Molecular linkage map of Einkorn wheat: mapping of storage-protein and soft-glume genes and bread-making quality QTLs.

Two molecular maps of Triticum monococcum L were produced and integrated. The integrated map includes a total of 477 markers, 32 RFLPs, 438 AFLPs, one morphological (soft glume (Sog)) and six storage-protein markers, and covers 856 cM. The trait Sog with the recessive allele sog maps to linkage group 2S. Probably, this is the T. monococcum homologue of Tg and Tg2 in hexaploid and tetraploid wheats, respectively. Loci coding for seed storage proteins were allocated to chromosomes 1L (HMW GLU1,2 and Glu1), 1S (LMW GLU6,7, LMW GLU1-4, omega GLI1-4, gamma GLI5 and Gli-1) and 6L (alpha/beta GLI7-14). Parameters related to bread-making quality (SDS sedimentation volume, specific sedimentation volume (SSV) and total protein content) were studied in one of the two populations. A QTL that is consistently present across environments was detected for SDS sedimentation volume and for SSV. The position of the QTL on chromosome 1S was in close agreement with the map positions of storage-protein loci. A second QTL was mapped on chromosome 5. For protein content, two significant QTLs were mapped to linkage groups 1 and 5.

A molecular, isozyme and morphological map of the barley (Hordeum vulgare) genome.

A map of the barley genome consisting of 295 loci was constructed. These loci include 152 cDNA restriction fragment length polymorphism (RFLP), 114 genomic DNA RFLP, 14 random amplified polymorphic DNA (RAPD), five isozyme, two morphological, one disease resistance and seven specific amplicon polymorphism (SAP) markers. The RFLP-identified loci include 63 that were detected using cloned known function genes as probes. The map covers 1,250 centiMorgans (cM) with a 4.2 cM average distance between markers. The genetic lengths of the chromosomes range from 124 to 223 cM and are in approximate agreement with their physical lengths. The centromeres were localized to within a few markers on all of the barley chromosomes except chromosome 5. Telomeric regions were mapped for the short (plus) arms of chromosomes 1, 2 and 3 and the long (minus) arm of chromosomes 7.