Thinopyrum distichum addition lines: production,morphological and cytological characterisation of 11 disomic addition lines and stable addition-substitution line

Plants of the partial amphiploid Inia 66/Thinopyrum distichum (2n = 70)//Inia 66 (2n = 56) were used as male parents in crosses with the monosomic series in the common wheat cultivar Inia 66. The genome and homoeologous group of the monosomic used in the cross affected the distribution of chromosome number of the progeny plants in the F₂ and F₄. Meiosis in the pollen mother cells of the B₁F₇ partial amphiploids was not stable, and not different from that of the B₁F₁ in which univalents and multivalents were observed. Disomic addition lines were selected on the basis of morphology and meiotic stability in the F₂, F₄ and F₅. Eleven of the fourteen possible wheat-Th. distichum disomic addition lines were identified using chromosome C-band pattern, as well as size and arm ratio, as genetic markers. Addition of T. distichum chromosome J ^dll produced a phenotype indicating homoeology with wheat group-2 chromosomes. Clear indications of homoeology based on morphological characteristics were not obtained in any of the other addition lines, probably due to the mixed homoeology of the Th. distichum chromosomes relative to wheat. The addition lines were all susceptible to leaf rust, unlike the germplasm-line Indis which carries a leaf rust resistance gene on a translocation segment derived from Th. distichum. Instability of meiotic pairing was observed in all addition lines. The stability, or not, of progeny chromosome counts did not reflect the level of chromosome pairing instability in the parental plants. SDS-PAGE for gliadin-type seed proteins revealed two addition lines which expressed seed storage proteins uncommon to Inia 66 but typical of Th. distichum.     

Direct evidence for high level of autosyndetic pairing in hybrids of Thinopyrum intermedium and Th. ponticum with Triticum aestivum

 Genomic in situ hybridization (GISH) was used to distinguish autosyndetic from allosyndetic pairing in the hybrids of Thinopyrum intermedium and Th. ponticum with Triticum aestivum cv ‘Chinese Spring’ (CS). All hybrids showed high autosyndetic pairing frequencies among wheat chromosomes and among Thinopyrum chromosomes. The high autosyndetic pairing frequencies among wheat chromosomes in both hybrids suggested that Th. intermedium and Th. ponticum carry promoters for homoeologous chromosome pairing. The higher frequencies of autosyndetic pairing among Thinopyrum chromosomes than among wheat chromosomes in both hybrids indicated that the relationships among the three genomes of Th. intermedium and among the five genomes of Th. ponticum are closer than those among the three genomes of T. aestivum. Received: 19 September 1996 / Accepted: 18 April 1997     

Introduction of Thinopyrum intermedium ssp. trichophorum chromosomes to wheat by trigeneric hybridization involving Triticum,Secale and Thinopyrum genera

Main conclusion

Fluorescence in situ hybridization and molecular markers have confirmed that several chromosomes from Thinopyrum intermedium ssp. trichophorum have been added to a wheat background, which originated from a cross between a wheat– Thinopyrum partial amphiploid and triticale. The lines displayed blue grains and resistance to wheat stripe rust.

Thinopyrum intermedium has been used as a valuable resource for improving the disease resistance and yield potential of wheat. With the aim to transfer novel genetic variation from Th. intermedium species for sustainable wheat breeding, a new trigeneric hybrid was produced by crossing an octoploid wheat–Th. intermedium ssp. trichophorum partial amphiploid with hexaploid triticale. Fluorescence in situ hybridization (FISH) revealed that Thinopyrum chromosomes were transmitted preferably and the number of rye chromosomes tended to decrease gradually in the selfed derivatives of the trigeneric hybrids. Four stable wheat–Th. intermedium chromosome substitution, addition and translocation lines were selected, and a 2J^S addition line, two substitution lines of 4J^S(4B) and 4J(4B), and a small 4J.4B translocation line were identified by FISH and molecular markers. It was revealed that the gene(s) responsible for blue grains may located on the FL0.60–1.00 of long arm of Th. intermedium-derived 4J chromosome. Disease resistance screenings indicated that chromosomes 4J^S and 2J^S appear to enhance the resistance to stripe rust in the adult plant stage. The new germplasm with Th. intermedium introgression shows promise for utilization of Thinopyrum chromosome segments in future wheat improvement.

     

Molecular Cytogenetics of an Amphiploid Trigeneric Hybrid between Triticum durum, Thinopyrum distichum and Lophopyrum elongatum

In situ hybridization of total genomic DNA was used to analyselines derived from an amphiploid between tetraploid wheat,Triticumdurum Desf. (2n =4x =28), and the wheatgrassesThinopyrum distichum(Thunb.) A. Löve (2n =4x =28) andLophopyrum elongatum (Host)A. Löve (2n =2x =14). A range of chromosome numbers wasdetected, arising from loss or gain of chromosomes. Total genomicDNA probes fromThinopyrum species,L. elongatum andTriticum monococcumL. were able to discriminate chromosomes from the A and B genomesof tetraploid wheat and those of wheatgrass-origin. The methoddid not discriminate the two wheatgrass genomes, J and E, indicatingtheir close similarity. Chromosomal aberrations—includingtelocentric and ring chromosomes—were frequent. Distalinter-genomic translocations of parts of A and B genome chromosomearms, unusual in wheat itself, were more frequent than translocationsbetweenT. durum and wheatgrass.In situ hybridization of an rDNAprobe most frequently revealed four sites associated with secondaryconstrictions onT. durum chromosomes and four onTh. distichumorL. elongatum chromosomes, although there was variation inthe number of loci between and within plants. Within interphaseand prophase nuclei, the three genomes were not intermixed andoften lay in distinct sectors. Wheat; hybrids; Triticum ; Triticeae; evolution; introgression; nuclear architecture; rDNA; in situ hybridization     

Molecular cytogenetic evidence for a high level of chromosome pairing among different genomes in Triticum aestivum–Thinopyrum intermedium hybrids

Intergeneric hybrids (ABDJJ^sS genomes) were made between Triticum aestivum cv. Chinese Spring (CS) and Thinopyrum intermedium. Genomic in situ hybridization (GISH) using genomic DNA probes from Pseudoroegneria libanotica (Hackel) D.R. Dewey (genome S, 2n = 14) was used to study chromosome pairing among J, J^s, S and wheat ABD genomes in the hybrids. It was shown that in the hexaploid (ABDJJ^sS) hybrids, high pairing occurred among wheat chromosomes and among Thinopyrum chromosomes. A closer relationship was observed among the three genomes of Th. intermedium than among the three genomes of T. aestivum. It was further discerned that S genome chromosomes paired with J- and J^s-genome chromosomes at a high frequency. The frequency of heterologous pairing between S and J or S and J^s chromosomes was higher than those between J and J^s chromosomes, indicating that the S-genome was more closely related with these two genomes. Our results provided direct molecular cytogenetic evidence for the hypothesis that S-genome chromosomes are genetically similar to the J-genome chromosomes and, therefore, genetic exchange between these genomes is possible. The discovery of a close relationship among S, J and J^s genomes provides valuable markers for molecular cytogenetic analyses using S-genomic DNA probes in monitoring the transfer of useful traits from Thinopyrum species into wheat. Received: 23 August 2000 / Accepted: 5 September 2000     

Molecular cytogenetic characterization of two high protein wheat-<Emphasis Type="Italic">Thinopyrum intermedium</Emphasis> 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.     

Expression of Thinopyrum distichum NORs in wheat×Thinopyrum amphiploids and their backcross generations

Summary The C-banding pattern of the satellited chromosomes in Thinopyrum distichum and Triticum durum was established. Both T. durum and Th. distichum contained two pairs of satellited chromosomes, which could be distinguished from one another. In the amphiploids [T. durum×Th. distichum (2x=56)] and in the backcross T. durum/(T. durum×Th. distichum)², BC₁F₃, and BC₁F₅ (2n = 42) the satellite was visible on only 1B and 6B of T. durum. The vector pTa 71 containing the rRNA gene from wheat hybridized to two pairs of chromosomes (four hybridization sites) in T. durum and Th. distichum, to eight sites in the amphiploid hybrid (2n=56), and to six sites in the backcross populations BC₁F₁. BC₁F₃, and BC₁F₅ (2n=42). The two satellite pairs in Th. distichum could be distinguished by the chromosomal location of the rRNA site (median or subterminal) and by the centromere position. One copy of each pair was present in the BC₁F₁, but in the BC₁F₃ and BC₁F₅ populations the pair with the subterminal location of rRNA genes was absent. Silver nitrate staining indicated that the rRNA genes of T. durum did not completely suppress those of Th. distichum. The octoploid amphiploid (2n = 56) contained a maximum of four large and four small nucleoli and the hexaploid BC₁s (2n=42), four large and two small nucleoli.     

Resistance to eyespot of wheat, caused by Tapesia yallundae, derived from Thinopyrum intermedium homoeologous group 4 chromosome

Thinopyrum intermedium was identified previously as resistant to Tapesia yallundae, cause of eyespot of wheat. Using GUS-transformed isolates of T. yallundae as inoculum, we determined that wheat lines carrying Th. intermedium chromosome 4Ai#2 or the short arm of chromosome 4Ai#2 were as resistant to the pathogen as the eyespot-resistant wheat- Th. ponticum chromosome substitution line SS767 (PI 611939) and winter wheat cultivar Madsen, which carries gene Pch1 for eyespot resistance. Chromosome 4E from Th. elongatum and chromosome 4J from Th. bessarabicum did not confer resistance to T. yallundae. Genome-specific PCR primers confirmed the presence of Thinopyrum chromatin in these wheat- Thinopyrum lines. Genomic in situ hybridization using an St genomic probe from Pseudoroegneria strigosa demonstrated that chromosome 4Ai#2 belongs to the J^s genome of Thinopyrum. The eyespot resistance in the wheat- Th. intermedium lines is thus controlled by the short arm of this J^s chromosome. This is the first report of resistance to T. yallundae controlled by a J^s genome chromosome of Th. intermedium.     

An efficient Oligo-FISH painting system for revealing chromosome rearrangements and polyploidization in Triticeae

A chromosome-specific painting technique has been developed which combines the most recent approaches of the companion disciplines of molecular cytogenetics and genome research. We developed seven oligonucleotide (oligo) pools derivd from single-copy sequences on chromosomes 1 to 7 of barley (Hordeum vulgare L.) and corresponding collinear regions of wheat (Triticum aestivum L.). The seven groups of pooled oligos comprised between 10 986 and 12 496 45-bp monomers, and these then produced stable fluorescence in situ hybridization (FISH) signals on chromosomes of each linkage group of wheat and barley. The pooled oligo probes were applied to high-throughput karyotyping of the chromosomes of other Triticeae species in the genera Secale, Aegilops, Thinopyrum, and Dasypyrum, and the study also extended to some wheat-alien amphiploids and derived lines. We demonstrated that a complete set of whole-chromosome oligo painting probes facilitated the study of inter-species chromosome homologous relationships and visualized non-homologous chromosomal rearrangements in Triticeae species and some wheat-alien species derivatives. When combined with other non-denaturing FISH procedures using tandem-repeat oligos, the newly developed oligo painting techniques provide an efficient tool for the study of chromosome structure, organization, and evolution among any wild Triticeae species with non-sequenced genomes.     

Genomic constitution and variation in five partial amphiploids of wheat–<Emphasis Type="Italic">Thinopyrum intermedium</Emphasis> as revealed by GISH,multicolor GISH and seed storage protein analysis

Genomic in situ hybridization (GISH) and multicolor GISH (mcGISH) methodology were used to establish the cytogenetic constitution of five partial amphiploid lines obtained from wheat × Thinopyrum intermedium hybridizations. Line Zhong 1, 2n=52, contained 14 chromosomes from each of the wheat genomes plus ten Th. intermedium chromosomes, with one pair of A-genome chromosomes having a Th. intermedium chromosomal segment translocated to the short arm. Line Zhong 2, 2n=54, had intact ABD wheat genome chromosomes plus 12 Th. intermedium chromosomes. The multicolor GISH results, using different fluorochrome labeled Th. intermedium and the various diploid wheat genomic DNAs as probes, indicated that both Zhong 1 and Zhong 2 contained one pair of Th. intermedium chromosomes with a significant homology to the wheat D genome. High-molecular-weight (HMW) glutenin and gliadin analysis revealed that Zhong 1 and Zhong 2 had identical banding patterns that contained all of the wheat bands and a specific HMW band from Th. intermedium. Zhong 1 and Zhong 2 had good HMW subunits for wheat breeding. Zhong 3 and Zhong 5, both 2n=56, possessed no gross chromosomal aberrations or translocations that were detectable at the GISH level. Zhong 4 also had a chromosome number of 2n=56 and contained the complete wheat ABD-genome chromosomes plus 14 Th. intermedium chromosomes, with one pair of Th. intermedium chromosomes being markedly smaller. Multicolor GISH results indicated that Zhong 4 also contained two pairs of reciprocally translocated chromosomes involving the A and D genomes. Zhong 3, Zhong 4 and Zhong 5 contained a specific gliadin band from Th. intermedium. Based on the above data, it was concluded that inter-genomic transfer of chromosomal segments and/or sequence introgression had occurred in these newly synthesized partial amphiploids despite their diploid-like meiotic behavior and disomic inheritance.     

A study of modified forms of the Lr19 translocation of common wheat

 Following the induction of allosyndetic pairing between the Thinopyrum-derived Lr19 translocation in ‘Indis’ wheat and homoeologous wheat chromatin, eight suspected recombinants for the Lr19 region were recovered. These selections were characterised for marker loci that were previously used to construct a physical map of the Lr19 segment. At the same time near-isogenic lines were developed for some of the selected segments and tested for seedling leaf-rust resistance in order to confirm the presence of Lr19. It appeared that three of the four white-endosperm selections do not possess Lr19 and only one, 88M22-149, is a true Lr19 recombinant. The resistance gene in the three non-Lr19 selections resides on chromosome 6B, appears to derive from ‘Indis’, and was selected unintentionally during backcrossing. The pedigree of ‘Indis’ is suspect and it is believed that the Lr19 translocation in ‘Indis’ is in reality the Th. ponticum-derived (T4) segment rather than being of Th. distichum origin as was believed earlier. The white-endosperm recombinant, 88M22-149, retained the complete Lr19 resistance and was apparently re-located to chromosome arm 7BL in a double-crossover event. 88M22-149 has lost the Sd1 gene and often shows strong self-elimination in translocation heterozygotes. This effect may result from additional gametocidal loci or from an altered chromosome structure following re-location of the segment. 88M22-149 in fact contains a duplicated region involving the Wsp-B1 locus. Three selections had partially white endosperms and expressed Lr19 and other Thinopyrum marker alleles. Polymorphisms for the available markers confirmed that the translocated segment in at least one of them had been shortened through recombination with chromosome arm 7DL. Further markers need to be studied in order to determine whether the translocation in the remaining two partially white recombinants had also undergone recombination with wheat. The eighth selection has yellow endosperm and appears to self-eliminate in certain translocation heterozygotes. No evidence of recombination could be found with the markers used. If the latter selections are in fact recombinants they may prove useful in attempts to unravel the complex segregation distortion mechanism. Received: 8 August 1996 / Accepted: 10 January 1997     

Chromosomes of Australian lygosomine skinks (Lacertilia: Scincidae)

The karyotypes of 25 species from the scincid genera Egernia, Corucia and Tiliqua have been investigated using C-banding, silver staining of nucleolar organiser regions (NORs) and Hoechst 33258 induced condensation inhibition. At least one member from each of the species groups of Egernia recognised by Storr et al. (1981) was studied. The three genera have very similar conventionally stained karyotypes of 32 chromosomes. Some species show departures from this basic karyotype but these are due to additions of C-band positive material. Silver stained. NOR patterns are variable but most species have a silver staining site on a pair of larger microchromosomes. All specimens studied except one have a proximal C-band on the acrocentric ninth pair, which shows failure to condense following treatment with the fluorochrome Hoechst 33258. Heterogamety was not observed in any species. Mabuya multifasciata, proposed as a relative of the Egernia group, while having 32 chromosomes does not share the C-band marker on pair nine, unique to the Egernia group. Tribolonotus gracilis, sometimes allied with the Egernia group, has 32 chromosomes and a similiar karyotype, but prominent procentric C-bands on all chromosome pairs obscure the detection of the proximal C-band marker on pair nine.     

Chromosome markers in Mus musculus: Differences in C-banding between the subspecies M. m. musculus and M. m. molossinus

Quinacrine (Q-band) and centromeric heterochromatin (C-band) patterns of metaphase chromosomes of two subspecies of Mus musculus were compared. M. m. musculus (the laboratory mouse) and M. m. molossinus (a subspecies from Southeast Asia) had similar Q-band patterns along the length of the chromosomes, but differences were observed in the centromeric region of some chromosomes. The two subspecies had very different distributions of C-band material. Antibodies to 5-methylcytosine were bound to regions of the chromosome corresponding to the C-bands in each animal. These findings support the idea that satellite DNA, which is concentrated in the C-band region, changes more quickly than bulk DNA. The interfertility of these two subspecies permits the development of a musculus strain carrying normal marker chromosomes for genetic studies.     

Cloning of resistance gene analogs located on the alien chromosome in an addition line of wheat-Thinopyrum intermedium

Homology-based gene/gene-analog cloning method has been extensively applied in isolation of RGAs (resistance gene analogs) in various plant species. However, serious interference of sequences on homoeologous chromosomes in polyploidy species usually occurred when cloning RGAs in a specific chromosome. In this research, the techniques of chromosome microdissection combined with homology-based cloning were used to clone RGAs from a specific chromosome of Wheat-Thinopyrum alien addition line TAi-27, which was derived from common wheat and Thinopyrum intermedium with a pair of chromosomes from Th. intermedium. The alien chromosomes carry genes for resistance to BYDV. The alien chromosome in TAi-27 was isolated by a glass needle and digested with proteinase K. The DNA of the alien chromosome was amplified by two rounds of Sau3A linker adaptor-mediated PCR. RGAs were amplified by PCR with the degenerated primers designed based on conserved domains of published resistance genes (R genes) by using the alien chromosome DNA, genomic DNA and cDNA of Th. intermedium, TAi-27 and 3B-2 (a parent of TAi-27) as templates. A total of seven RGAs were obtained and sequenced. Of which, a constitutively expressed single-copy NBS-LRR type RGA ACR3 was amplified from the dissected alien chromosome of TAi-27, TcDR2 and TcDR3 were from cDNA of Th. intermedium, AcDR3 was from cDNA of TAi-27, FcDR2 was from cDNA of 3B-2, AR2 was from genomic DNA of TAi-27 and TR2 was from genomic DNA of Th. intermedium. Sequence homology analyses showed that the above RGAs were highly homologous with known resistance genes or resistance gene analogs and belonged to NBS-LRR type of R genes. ACR3 was recovered by PCR from genomic DNA and cDNA of Th. intermedium and TAi-27, but not from 3B-2. Southern hybridization using the digested genomic DNA of Th. intermedium, TAi-27 and 3B-2 as the template and ACR3 as the probe showed that there is only one copy of ACR3 in the genome of Th. intermedium and TAi-27, but it is absent in 3B-2. The ACR3 could be used as a specific probe of the R gene on the alien chromosome of TAi-27. Results of Northern hybridization suggested that ACR3 was constitutively expressed in Th. intermedium and TAi-27, but not 3B-2, and expressed higher in leaves than in roots. This research demonstrated a new way to clone RGAs located on a specific chromosome. The information reported here should be useful to understand the resistance mechanism of, and to clone resistant genes from, the alien chromosome in TAi-27.     

Karyomorphological parameters and C-band distribution suggest phyletic relationships within the subtribeLimodorinae (Orchidaceae)

Karyotype attributes and heterochromatin distribution were used to characterize fourteen taxa of the subtribeLimodorinae (Orchidaceae). The karyotypes were established using morphometrical parameters following Feulgen staining and C-banding. No significant differences in heterochromatin content were found between specimens collected from various sites. Four species of theEpipactis helleborine group possess some chromosome pairs with quite similar heterochromatin patterns; some differences were found inE. distans with respect to other species of this group.Epipactis palustris differed significantly from otherEpipactis species in its different karyotype and its numerous terminal C-bands. The largest differences from the other genera were shown inLimodorum as far as karyomorphology and heterochromatin patterns were concerned. C-band distribution indicated similarity among non-homologous chromosomes, supporting a possible palaeo-polyploid origin for theCephalanthera andEpipactis karyotypes.     

Giemsa C-banding patterns in Aedes (Stegomyia) mosquitoes

Chromosomes from gonads of 14–24 h old pupae of nine species of Stegomyia mosquitoes have been examined using the Giemsa C-banding technique. The species studied were Aedes albopictus, A. polynesienis, A. scutellaris, A. alcasidi, A. seatoi, A. pseudalbopictus, A. melallicus, A. annandalei and A. vittatus. The diploid chromosome number of all species is six. All species possess C-bands in the centromeric regions of each of the three pairs of chromosomes. Besides, an intercalary C-band is present on the female determining (=m) chromosome but absent from the male—determining (= M) chromosome of all species except A. vittatus. In A. vittatus, the m and M chromosomes possess a terminal C-band. Thus, the nine species of Aedes analysed showed two distinct patterns of C-banding. —The evolution of heterochromatin patterns in various species is also discussed. The Giemsa C-banding technique should prove useful in studies of chromosomal speciation in culicine mosquitoes.     

Molecular cytogenetic characterization of four partial wheat-Thinopyrum ponticum amphiploids and their reactions to Fusarium head blight, tan spot, and Stagonospora nodorum blotch

Four wheat (Triticum aestivum L.)-Thinopyrum ponticum derivatives SS5 (PI604926), SS156 (PI604947), SS363 (PI604970), and SS660 (PI604879), were identified as resistant to Fusarium head blight (FHB), a serious fungal disease of wheat worldwide. Seedling reactions to tan spot and Stagonospora nodorum blotch (SNB), two important foliar diseases of wheat, suggest that these four derivatives are resistant to tan spot and two of them (SS5 and SS156) are resistant to SNB. Fluorescent genomic in situ hybridization (FGISH) patterns of mitotic chromosomes indicate that these four derivatives are partial wheat-Th. ponticum amphiploids, each with a total of 56 chromosomes, though with different amounts of Th. ponticum chromatin. These four amphiploids were hybridized with each other to determine homology between the Th. ponticum genomes in each of the amphiploids. Analysis of chromosome pairing in the F₁ hybrids using FGISH suggests that each amphiploid carries a similar set of Th. ponticum chromosomes. These wheat-Th. ponticum amphiploids represent a potential novel source of resistance to FHB, tan spot, and SNB for wheat breeding.     

Regularities and restrictions governing C-band variation in acridoid grasshoppers

C-band patterns have been analysed in embryonic neuroblast chromosomes of 23 Australian species of acridoids. All of them showed paracentromeric C-bands but these varied considerably in size both within and between species. Many of them also showed interstitial C-bands in from 1–5 members of the haploid complement and in two cases (Atractomorpha similis and Genus nov. 95 ochracea) larger numbers of interstitial bands were present. Terminal C-bands were the least common though again when present they could be found in 1–6 members of the complement except in the cases of A. similis and Genus nov. 95 ochracea where still larger numbers occur. In 5 of the 23 species the megameric chromosome pair was distinctively C-banded. The B-chromosomes found in 3 of the species were also strikingly different in C-band characteristics compared to the standard A-chromosomes. Differences in the number of very small chromosomes present in different species clearly cannot be explained in terms of differences in their C-band content. Neither are differences in genome size simply related to differences in the total amount of C-band material indicating that changes in the size of the genome in this group have involved alterations in both eu and heterochromatin content. Finally similar amounts of C-band material may be distributed throughout the complement in very different ways in different species.To Hans Bauer with respect and affection on the occasion of his 75th birthday.     

Development and utilization of new sequenced characterized amplified region markers specific for E genome of Thinopyrum

Species containing E genome of Thinopyrum offered potential to increase the genetic variability and desirable characters for wheat improvement. However, E genome specific marker was rare. The objective of the present report was to develop and identify sequenced characterized amplified region (SCAR) markers that can be used in detecting E chromosome in wheat background for breeding purpose. Total 280 random amplified polymorphic DNA (RAPD) primers were amplified for seeking of E genome specific fragments by using the genomic DNA of Thinopyrum elongatum and wheat controls as templates. As a result, six RAPD fragments specific for E genome were found and cloned, and then were converted to SCAR markers. The usability of these markers was validated using a number of Egenome-containing species and wheat as controls. These markers were subsequently located on E chromosomes using specific PCR and fluorescence in situ hybridization (FISH). SCAR markers developed in this research could be used in molecular marker assisted selection of wheat breeding with Thinopyrum chromatin introgressions.