PCR-based Landmark Unique Gene Research Articles

Cytological and molecular characterization of wheat lines carrying leaf rust and stem rust resistance genes Lr24 and Sr24

Previous studies showed that Australian wheat cultivars Janz and Sunco carry leaf rust and stem rust resistance genes Lr24 and Sr24 derived from Thinopyrum ponticum chromosome arm 3AgL. However, the size of the alien segments carrying Lr24 and Sr24 in the lines were not determined. In this study, we used non-denaturing fluorescence in situ hybridization (ND-FISH), genomic in situ hybridization (GISH), and PCR-based landmark unique gene (PLUG) markers to visualize the alien segments in Janz and Sunco, and further compared them with the segments in US cultivars Agent and Amigo. The fraction length (FL) of the alien translocation in Agent was 0.70–1.00, whereas those in Janz, Sunco, and Amigo were smaller, at FL 0.85–1.00. It was deduced that the alien gene RAg encoding for red grain color and rust resistance genes Lr24 and Sr24 on chromosome arm 3AgL were in bins of FL 0.70–0.85 and 0.85–1.00, respectively. We retrieved and extracted nucleotide-binding site-leucine-rich repeat (NBS-LRR) receptor genes corresponding to the region of Lr24 and Sr24 on chromosomes 3E, and 3J, 3Js and 3St from the reference genome sequences of Th. elongatum and Th. intermedium, respectively. A set of molecular markers developed for Lr24 and Sr24 from those extracted NBS-LRR genes will provide valuable information for fine mapping and cloning of these genes.

Molecular and Cytogenetic Identification of Stem Rust Resistant Wheat-Thinopyrum intermedium Introgression Lines.

Thinopyrum intermedium (JJJsJsStSt, 2n = 6x = 42), a wild relative of common wheat, possesses many desirable agronomic genes for wheat improvement. The production of wheat-Thinopyrum intermedium introgression lines is a key step for transferring these beneficial genes into wheat. In this study, we characterized three wheat-Thinopyrum intermedium introgression lines TA3681, TA5566, and TA5567 using non-denaturing fluorescence in situ hybridization, genomic in situ hybridization, PCR-based landmark unique gene, and intron targeting markers. Our results showed that TA3681 is a wheat-Thinopyrum intermedium 1Stdisomic addition line, TA5566 is a wheat-Thinopyrum intermedium non-Robertsonian translocation line carrying two pairs of 3A-7Js translocation chromosomes, and that TA5567 is a wheat-Thinopyrum intermedium non-Robertsonian translocation line carrying a pair of 3A-7Js translocation chromosomes. We developed 13, 36, and 15 Thinopyrum intermedium chromosome-specific markers for detecting the introgressed Thinopyrum chromosomes in TA3681, TA5566, and TA5567, respectively. Stem rust assessment revealed that TA3681 exhibited ahigh level of seedling resistance to Chinese-prevalent Puccinia graminis f. sp. tritici pathotypes, and both TA5566 and TA5567 were highly resistant to Australian P. graminis f. sp. tritici pathotypes, indicating that Thinopyrum intermedium chromosomes 1St and 7Js might carry new stem rust resistance genes. Therefore, the new identified introgression lines may be useful for improving wheat stem rust resistance.

Identification and Evaluation of Wheat-Aegilops bicornis Lines with Resistance to Powdery Mildew and Stripe Rust.

Wheat pathogens, especially those causing powdery mildew and stripe rust, seriously threaten yield worldwide. Utilizing newly identified disease resistance genes from wheat relatives is an effective strategy to minimize disease damage. In this study, chromosome-specific molecular markers for the 3Sb and 7Sb chromosomes of Aegilops bicornis were developed using PCR-based landmark unique gene primers for screening wheat-A. bicornis progenies. Fluorescence in situ hybridization (FISH) was performed to further identify wheat-A. bicornis progenies using oligonucleotides probes Oligo-pSc119.2-1, Oligo-pTa535-1, and Oligo-(GAA)8. After establishing A. bicornis 3Sb and 7Sb chromosome-specific FISH markers, Holdfast (common wheat)-A. bicornis 3Sb addition, 7Sb addition, 3Sb(3A) substitution, 3Sb(3B) substitution, 3Sb(3D) substitution, 7Sb(7A) substitution, and 7Sb(7B) substitution lines were identified by the molecular and cytological markers. Stripe rust and powdery mildew resistance, along with agronomic traits, were investigated to evaluate the breeding potential of these lines. Holdfast and Holdfast-A. bicornis progenies were all highly resistant to stripe rust, indicating that the stripe rust resistance might derive from Holdfast. However, Holdfast-A. bicornis 3Sb addition, 3Sb(3A) substitution, 3Sb(3B) substitution, and 3Sb(3D) substitution lines showed high resistance to powdery mildew while Holdfast was highly susceptible, indicating that chromosome 3Sb of A. bicornis carries previously unknown powdery mildew resistance gene(s). Additionally, the transfer of the 3Sb chromosome from A. bicornis to wheat significantly increased tiller number, but chromosome 7Sb has a negative effect on agronomic traits. Therefore, wheat germplasm containing A. bicornis chromosome 3Sb has potential to contribute to improving powdery mildew resistance and tiller number during wheat breeding.

Identification of a Wheat-Psathyrostachys huashanica 7Ns Ditelosomic Addition Line Conferring Early Maturation by Cytological Analysis and Newly Developed Molecular and FISH Markers.

Early maturation is an important objective in wheat breeding programs that could facilitate multiple-cropping systems, decrease disaster- and disease-related losses, ensure stable wheat production, and increase economic benefits. Exploitation of novel germplasm from wild relatives of wheat is an effective means of breeding for early maturity. Psathyrostachys huashanica Keng f. ex P. C. KUO (2n=2x=14, NsNs) is a promising source of useful genes for wheat genetic improvement. In this study, we characterized a novel wheat-P. huashanica line, DT23, derived from distant hybridization between common wheat and P. huashanica. Fluorescence in situ hybridization (FISH) and sequential genomic in situ hybridization (GISH) analyses indicated that DT23 is a stable wheat-P. huashanica ditelosomic addition line. FISH painting and PCR-based landmark unique gene markers analyses further revealed that DT23 is a wheat-P. huashanica 7Ns ditelosomic addition line. Observation of spike differentiation and the growth period revealed that DT23 exhibited earlier maturation than the wheat parents. This is the first report of new earliness per se (Eps) gene(s) probably associated with a group 7 chromosome of P. huashanica. Based on specific locus-amplified fragment sequencing technology, 45 new specific molecular markers and 19 specific FISH probes were developed for the P. huashanica 7Ns chromosome. Marker validation analyses revealed that two specific markers distinguished the Ns genome chromosomes of P. huashanica and the chromosomes of other wheat-related species. These newly developed FISH probes specifically detected Ns genome chromosomes of P. huashanica in the wheat background. The DT23 line will be useful for breeding early maturing wheat. The specific markers and FISH probes developed in this study can be used to detect and trace P. huashanica chromosomes and chromosomal segments carrying elite genes in diverse materials.

Molecular cytogenetics for a wheat\u2013Aegilops geniculata 3Mg alien addition line with resistance to stripe rust and powdery mildew

BackgroundAegilops geniculata Roth is closely related to common wheat (Triticum aestivum L.) and is a valuable genetic resource for improvement of wheat.ResultsIn this study, the W19513 line was derived from the BC1F10 progeny of a cross between wheat ‘Chinese Spring’ and Ae. geniculata SY159. Cytological examination showed that W19513 contained 44 chromosomes. Twenty-two bivalents were formed at the first meiotic metaphase I in the pollen mother cellsand the chromosomes were evenly distributed to opposite poles at meiotic anaphase I. Genomic in situ hybridization demonstrated that W19513 carried a pair of alien chromosomes from the M genome. Fluorescence in situ hybridization confirmed detection of variation in chromosomes 4A and 6B. Functional molecular marker analysis using expressed sequence tag–sequence-tagged site and PCR-based landmark unique gene primers revealed that the alien gene belonged to the third homologous group. The marker analysis confirmed that the alien chromosome pair was 3Mg. In addition, to further explore the molecular marker specificity of chromosome 3Mg, based on the specific locus amplified fragment sequencing technique, molecular markers specific for W19513 were developed with efficiencies of up to 47.66%. The W19513 line was inoculated with the physiological race E09 of powdery mildew (Blumeria graminis f. sp. tritici) at the seedling stage and showed moderate resistance. Field inoculation with a mixture of the races CYR31, CYR32, CYR33, and CYR34 of the stripe rust fungus (Puccinia striiformis f. sp. triticii) revealed that the line W19513 showed strong resistance.ConclusionsThis study provides a foundation for use of the line W19513 in future genetic research and wheat improvement.

Production and characterization of a disomic 1M/1D Triticum aestivum-Aegilops comosa substitution line.

The online version contains supplementary material available at 10.1007/s11032-021-01207-2.

Physical mapping of chromosome 7J and a purple coleoptile gene from Thinopyrum intermedium in the common wheat background.

A physical map of Thinopyrum intermedium chromosome 7J was constructed using translocation mapping, and a new seedling purple coleoptile gene was mapped to the bin of FL 0.35-0.63 of 7JS. Thinopyrum intermedium (2n = 6x = 42, JJJsJsStSt), a wild relative of common wheat, harbors numerous beneficial genes for wheat improvement. Previous studies showed that wheat-Th. intermedium partial amphiploid TAF46 and its derived addition line L1 had a purple coleoptile, which was derived from Th. intermedium chromosome 7J. To identify and physically map the purple coleoptile gene, 12 wheat-Th. intermedium 7J translocation lines were analyzed by sequential multicolor fluorescence in situ hybridization (mc-FISH), PCR-based landmark unique gene (PLUG) and intron targeting (IT) markers. A physical map of the 7J chromosome was constructed, consisting of eight chromosomal bins with 89 markers. Seedling evaluation of the coleoptile colors of all tested materials indicated that the purple coleoptile gene was located to the bin with a fraction length (FL) of 0.35-0.63 on chromosome 7JS. Furthermore, based on the syntenic relationships between Th. intermedium and wheat chromosomes, we developed a new chromosome 7J-specific EST-PCR marker from the chromosomal region corresponding to the purple coleoptile gene through the Triticeae multi-omics database. The approach of designing chromosome-specific markers has facilitated fine mapping of the Thinopyrum-specific purple coleoptile gene, and these translocation lines will be valuable for studying the function of the purple coleoptile gene in anthocyanin biosynthesis.

Development and characterization of a wheat–Leymus mollis Lm#7Ns disomic addition line with resistance to stripe rust

A new wheat–Leymus mollis (2n = 4x = 28, NsNsXmXm) disomic addition line M11003-3-1-12-10 (M10) was developed and selected from the BC1F5 progeny of a cross between wheat cv. 7182 and octoploid Tritileymus M47 (2n = 8x = 56, AABBDDNsNs). Cytological observations showed that M10 contained 44 chromosomes and formed 22 bivalents at meiotic metaphase I. The sequential fluorescence in situ hybridization–genomic in situ hybridization (FISH–GISH) analysis using the oligonucleotide Oligo-pSc119.2, Oligo-pTa535 and L. mollis genomic DNA as probes revealed that M10 carried a pair of L. mollis chromosomes and a whole set of 42 wheat chromosomes. Two EST–STS markers and six PLUG (PCR-based Landmark Unique Gene) markers confirmed that the added L. mollis chromosomes in M10 belonged to homoeologous group 7 and were designated as Lm#7Ns, so M10 was deduced to be a wheat–L. mollis Lm#7Ns disomic addition line. Field disease evaluation demonstrated that M10 showed a high level of resistance to stripe rust at the adult stage and the resistance could be ultimately traced to the L. mollis. Consequently, the introduced Lm#7Ns chromosomes were responsible for the stripe rust resistance. The novel disomic addition line M10 could be exploited as an important genetic resource for wheat resistance breeding.

Characterization and Evaluation of Resistance to Powdery Mildew of Wheat-Aegilops geniculata Roth 7Mg (7A) Alien Disomic Substitution Line W16998.

Aegilops geniculata Roth has been used as a donor of disease-resistance genes, to enrich the gene pool for wheat (Triticum aestivum) improvement through distant hybridization. In this study, the wheat–Ae. geniculata alien disomic substitution line W16998 was obtained from the BC1F8 progeny of a cross between the common wheat ‘Chinese Spring’ (CS) and Ae. geniculata Roth (serial number: SY159//CS). This line was identified using cytogenetic techniques, analysis of genomic in situ hybridization (GISH), functional molecular markers (Expressed sequence tag-sequence-tagged site (EST–STS) and PCR-based landmark unique gene (PLUG), fluorescence in situ hybridization (FISH), sequential fluorescence in situ hybridization–genomic in situ hybridization (sequential FISH–GISH), and assessment of agronomic traits and powdery mildew resistance. During the anaphase of meiosis, these were evenly distributed on both sides of the equatorial plate, and they exhibited high cytological stability during the meiotic metaphase and anaphase. GISH analysis indicated that W16998 contained a pair of Ae. geniculata alien chromosomes and 40 common wheat chromosomes. One EST–STS marker and seven PLUG marker results showed that the introduced chromosomes of Ae. geniculata belonged to homoeologous group 7. Nullisomic–tetrasomic analyses suggested that the common wheat chromosome, 7A, was absent in W16998. FISH and sequential FISH–GISH analyses confirmed that the introduced Ae. geniculata chromosome was 7Mg. Therefore, W16998 was a wheat–Ae. geniculata 7Mg (7A) alien disomic substitution line. Inoculation of isolate E09 (Blumeria graminis f. sp. tritici) in the seedling stage showed that SY159 and W16998 were resistant to powdery mildew, indeed nearly immune, whereas CS was highly susceptible. Compared to CS, W16998 exhibited increased grain weight and more spikelets, and a greater number of superior agronomic traits. Consequently, W16998 was potentially useful. Germplasms transfer new disease-resistance genes and prominent agronomic traits into common wheat, giving the latter some fine properties for breeding.

Identification and characterization of a new stripe rust resistance gene Yr83 on rye chromosome 6R in wheat.

A physical map of Secale cereale chromosome 6R was constructed using deletion mapping, and a new stripe rust resistance gene Yr83 was mapped to the deletion bin of FL 0.73-1.00 of 6RL. Rye (Secale cereale L., RR) possesses valuable genes for wheat improvement. In the current study, we report a resistance gene conferring stripe rust resistance effective from seedling to adult plant stages located on chromosome 6R. This chromosome was derived from triticale line T-701 and also carries highly effective resistance to the cereal cyst nematode species Heterodera avenae Woll. A wheat-rye 6R(6D) disomic substitution line exhibited high levels of seedling resistance to Australian pathotypes of the stripe rust (Puccinia striiformis f. sp. tritici; Pst) pathogen and showed an even greater resistance to the Chinese Pst pathotypes in the field. Ten chromosome 6R deletion lines and five wheat-rye 6R translocation lines were developed earlier in the attempt to transfer the nematode resistance gene to wheat and used herein to map the stripe rust resistance gene. These lines were subsequently characterized by sequential multicolor fluorescence in situ hybridization (mc-FISH), genomic in situ hybridization (GISH), mc-GISH, PCR-based landmark unique gene (PLUG), and chromosome 6R-specific length amplified fragment sequencing (SLAF-Seq) marker analyses to physically map the stripe rust resistance gene. The new stripe rust resistance locus was located in a chromosomal bin with fraction length (FL) 0.73-1.00 on 6RL and was named Yr83. A wheat-rye translocation line T6RL (#5) carrying the stripe rust resistance gene will be useful as a new germplasm in breeding for resistance.

Comparative FISH and molecular identification of new stripe rust resistant wheat-Thinopyrum intermedium ssp. trichophorum introgression lines

Thinopyrum intermedium (2n = 6x = 42, JJJsJsStSt) has been hybridized extensively with common wheat and has proven to be a valuable germplasm source for improving disease resistance, quality attributes, and yield potential in wheat. We characterized new disease resistant wheat-Th. intermedium derivatives A1082 and A5-5 using sequential multi-color fluorescence in situ hybridization (mc-FISH), genomic in situ hybridization (GISH), PCR-based landmark unique gene (PLUG) and intron targeting (IT) markers. A1082 was identified as a wheat-Th. intermedium 3J disomic addition line, and A5-5 was a T4BS·5JsL homozygous Robertsonian translocation line. Seventy-one and 106 pairs of primers amplified Th. intermedium-specific bands allowing chromosomes 3J and 5Js to be tracked, respectively. A new oligonucleotide probe, Oligo-6H-2-100, was developed for FISH labeling of the subterminal region of the long arm of chromosome 5Js. Both lines were highly resistant to stripe rust pathogen races prevalent in Chinese field screening nurseries. A5-5 also displayed a significant increase in tiller number compared to its wheat parent. The new lines can be exploited as useful germplasms for wheat improvement.

Disomic chromosome addition from Thinopyrum intermedium to bread wheat appears to confer stripe rust resistance

A wheat line 14-569 developed from a hybrid of common wheat cv. Chuannong 10 with the wheat–Thinopyrum intermedium partial amphiploid TAI7045 exhibits stable double spikelets per node character in the lower middle portion of a spike, is highly resistant to stripe rust, and produces kernels with a higher 1000-kernel weight than those of its wheat parent. Chromosome counting and an analysis of meiosis demonstrated that line 14-569 is a disomic addition line with 2n = 44, 22II. Sequential genomic in situ hybridization and non-denaturing fluorescence in situ hybridization analyses indicated that one pair of St-genomic chromosomes from Th. intermedium was added to the wheat complement and that the wheat complement included one pair of 1RS/1BL wheat–rye translocation chromosomes. PCR-based landmark unique gene molecular marker analysis revealed that the added pair of chromosomes comprised 3St chromosomes, which suggested that the added 3St chromosomes from Th. intermedium might carry the genes to control the high yield-related characters and stripe rust resistance. This addition line will potentially be useful resource for improvements in wheat yields and stripe rust resistance.

Molecular markers and cytogenetics to characterize a wheat-Dasypyrum villosum 3V (3D) substitution line conferring resistance to stripe rust.

Dasypyrum villosum has been used as a valuable gene resource for disease resistances, yield increase and quality improvement in wheat. A novel wheat-D. villosum alien introgression line CD-3 was generated through hybridization between the common wheat Chinese Spring (CS) and a CS- D. villosum 3V addition line having considerably high stripe rust resistance, which enable the characterization of a potential new stripe rust resistance gene (s) derived from D. villosum. The results of non-denaturing fluorescent in situ hybridization (ND-FISH) showed that CD-3 contained 42 chromosomes, including a 3V chromosome pair, and the absence of both of the 3D chromosomes. PCR-based Landmark Unique Gene (PLUG) molecular marker analysis supported results from the FISH analysis, revealing CD-3 was a wheat-D. villosum 3V (3D) disomic substitution line. Resistant test of stripe rust on 52 plants of F2 generation (CD-3/CS), CD-3, CS and D.villosum have been conducted at seedling stage. 7 plants of F2 generation possessing two 3V chromosomes exhibited high resistance to stripe rust as CD-3 and D.villosum, 10 plants carrying one 3V chromosome and 35 plants without 3V chromosome were susceptive to stripe rust as CS. The result implied the high stripe rust resistance of CD-3 should be controlled by recessive gene(s) originating from D.villosum. To rapidly detect chromosome 3V in the genetic background of wheat, we developed a novel Sequence Characterized Amplified Region (SCAR) marker specific for 3V chromosome based on the sequence of a grain size-related gene DvGS5 in D. villosum, an orthologue of TaGS5 from wheat. The SCAR marker was designated DvGS5-1443, which could successfully amplify a unique 3V-specific fragment in CD-3 and D. villosum, suggesting that this SCAR marker could facilitate targeting the chromosome 3V in the genetic background of wheat for wheat improvement.

Molecular cytogenetic identification of a wheat - Thinopyrum ponticum substitution line with stripe rust resistance.

Thinopyrum ponticum (Th. ponticum) (2n = 10x = 70) is an important breeding material with excellent resistance and stress tolerance. In this study, we characterized the derivative line CH1113-B13-1-1-2-1 (CH1113-B13) through cytological, morphological, genomic in situ hybridization (GISH), fluorescence in situ hybridization (FISH), expressed sequence tag (EST), and PCR-based landmark unique gene (PLUG) marker analysis. The GISH analysis revealed that CH1113-B13 contained 20 pairs of common wheat chromosomes and one pair of JSt genomic chromosomes. Linkage analysis of Th. ponticum using seven EST and seven PLUG markers indicated that the pair of alien chromosomes belonged to the seventh homeologous group. Nulli-tetrasomic and FISH analysis revealed that wheat 7B chromosomes were absent in CH1113-B13; thus, CH1113-B13 was identified as a 7JSt (7B) substitution line. Finally, adult-stage CH1113-B13 exhibited immunity to wheat stripe rust. This substitution line is therefore a promising germplasm resource for wheat breeding.

A novel compensating wheat-Thinopyrum elongatum Robertsonian translocation line with a positive effect on flour quality.

Wheat flours are used to produce bread, pasta, breakfast cereals, and biscuits; the various properties of these end-products are attributed to the gluten content, produced as seed storage proteins in the wheat endosperm. Thus, genes encoding gluten protein are major targets of wheat breeders aiming to improve the various properties of wheat flour. Here, we describe a novel compensating wheat–Thinopyrum elongatum Robertsonian translocation (T1AS.1EL) line involving the short arm of wheat chromosome 1A (1AS) and the long arm of Th. elongatum chromosome 1E (1EL); we developed this line through centric breakage-fusion. Compared to the common wheat cultivars Chinese Spring and Norin 61, we detected two additional 1EL-derived high-molecular-weight glutenin subunits (HMW-GSs) in the T1AS.1EL plants. Based on the results of an SDS-sedimentation volume to estimate the gluten strength of T1AS.1EL-derived flour, we predict that T1AS.1EL-derived flour is better suited to bread-making than Chinese Spring- and Norin 61-derived flour and that this is because of its greater gluten diversity. Also, we were able to assign 33 of 121 wheat PCR-based Landmark Unique Gene markers to chromosome 1E of Th. elongatum. These markers can now be used for further chromosome engineering of the Th. elongatum segment of T1AS.1EL.

Development and molecular cytogenetic identification of a new wheat–Leymus mollis Lm#6Ns disomic addition line

AbstractWe developed a new disomic addition line M11028‐1‐1‐5 (2n = 44 = 21” + 1”) from a cross between wheat cv. ‘7182’ and octoploid Tritileymus M47 (2n = 8x = 56, AABBDDNsNs). Cytological observations demonstrated that M11028‐1‐1‐5 contained 44 chromosomes and formed 22 bivalents during meiotic metaphase I. The genomic in situ hybridization (GISH) investigations showed this line contained 42 wheat chromosomes and a pair of L. mollis chromosomes. SSR, EST and PCR‐based landmark unique gene (PLUG) markers were screened to determine the homoeologous relationships of the introduced L. mollis chromosomes in wheat background. Nine markers, i.e. Xwmc256, Xgpw312, Swes123, CD452568, BF483643, BQ169205, TNAC1748, TNAC1751 and TNAC1752, all of which were located on the homoeologous group 6 chromosomes of common wheat, amplified bands unique to L. mollis in M11028‐1‐1‐5. Gliadin analysis also confirmed that the added chromosomes in M11028‐1‐1‐5 were correlated with the sixth group chromosome. This indicated that M11028‐1‐1‐5 contained a pair of introduced L. mollis chromosome belonging to homoeologous group 6, which we designated it as Lm#6Ns disomic addition line. This is the first report of a common wheat–L. mollis disomic addition line.

Molecular cytogenetic identification of a wheat–Aegilops geniculata Roth 7Mg disomic addition line with powdery mildew resistance

Aegilops geniculata Roth is an important germplasm resource for the transfer of beneficial genes into common wheat (Triticum aestivum L.). A new disomic addition line NA0973-5-4-1-2-9-1 was developed from the BC1F6 progeny of the cross wheat cv. Chinese Spring (CS)/Ae. geniculata SY159//CS. We characterized this new line by morphological and cytogenetic identification, analysis of functional molecular markers, genomic in situ hybridization (GISH), fluorescence in situ hybridization (FISH), and disease resistance evaluation. Cytological observations suggested that NA0973-5-4-1-2-9-1 contained 44 chromosomes and formed 22 bivalents at meiotic metaphase I. The GISH investigations showed that the line contained 42 wheat chromosomes and a pair of Ae. geniculata chromosomes. EST-STS multiple loci markers and PLUG (PCR-based landmark unique gene) markers confirmed that the introduced Ae. geniculata chromosomes belonged to homoeologous group 7. FISH identification suggested that NA0973-5-4-1-2-9-1 possessed an additional pair of 7Mg chromosomes, and at the same time, there were structural differences in a pair of 6D chromosomes between NA0973-5-4-1-2-9-1 and TA7661 (CS-AEGEN DA 7Mg). After inoculation with powdery mildew (Blumeria graminis f. sp. tritici, Bgt) isolates E09, NA0973-5-4-1-2-9-1 exhibited a powdery mildew resistance infection type different from that of TA7661, and we conclude that the powdery mildew resistance of NA0973-5-4-1-2-9-1 originated from its parent Ae. geniculata SY159. Therefore, NA0973-5-4-1-2-9-1 can be used as a donor source for introducing novel disease resistance genes into wheat during breeding programs with the assistance of molecular and cytogenetic markers.

A Robertsonian translocation from Thinopyrum bessarabicum into bread wheat confers high iron and zinc contents

AbstractDevelopment of wheat–alien translocation lines has facilitated practical utilization of alien species in wheat improvement. The production of a compensating Triticum aestivum–Thinopyrum bessarabicum whole‐arm Robertsonian translocation (RobT) involving chromosomes 6D of wheat and 6Eb of Th. bessarabicum (2n = 2x = 14, EbEb) through the mechanism of centric breakage–fusion is reported here. An F2 population was derived from plants double‐monosomic for chromosome 6D and 6Eb from crosses between a DS6Eb(6D) substitution line and bread wheat cultivar ‘Roushan’ (2n = 6x = 42, AABBDD) as female parent. Eighty F2 genotypes (L1–L80) were screened for chromosome composition. Three PCR‐based Landmark Unique Gene (PLUG) markers specific to chromosomes 6D and 6Eb were used for screening the F2 plants. One plant with a T6EbS.6DL centric fusion (RobT) was identified. A homozygous translocation line with full fertility was recovered among F3 families and verified with genomic in situ hybridization (GISH). Grain micronutrient analysis showed that the DS6Eb(6D) substitution line and T6EbS.6DL stock have higher Fe and Zn contents than the recipient wheat cultivar ‘Roushan’.

Production of a novel Robertsonian translocation from Thinopyrum bessarabicum into bread wheat

Development of wheat-alien translocation lines will facilitate its practical utilization in wheat improvement. The objective of the present study was to produce compensating wheat--Thinopyrum bessarabicum whole arm Robertsonian translocations (RobTs) involving chromosomes 2B of wheat and 2E(b) of Th. bessarabicum through the mechanism of centric breakage-fusion. F2 population from crosses between DS2E(b)(2B) substitution line and bread wheat 'Roushan' (2n = 6x = 42, AABBDD) as female parent were made. Forty one F2 lines (L1 to L41) were screened for their chromosome composition. Three 2E(b) specific PCR-based Landmark Unique Gene (PLUG) markers were used for screening F2 progeny derived from plants double-monosomic for chromosome 2B and 2E(b). Two Rob Ts (-5%) were observed among F2 plants. Homozygous translocation (T2E(b)S.2BL) with good plant vigor and full fertility were selected from F3 families. The T2E(b)S.2BL stock has longer awn than that of its parents. It is cytogenetically stable, and may be useful in wheat improvement.

Development and Molecular Cytogenetic Identification of a Novel Wheat-Leymus mollis Lm#7Ns (7D) Disomic Substitution Line with Stripe Rust Resistance.

Leymus mollis (2n = 4x = 28, NsNsXmXm) possesses novel and important genes for resistance against multi-fungal diseases. The development of new wheat—L. mollis introgression lines is of great significance for wheat disease resistance breeding. M11003-3-1-15-8, a novel disomic substitution line of common wheat cv. 7182 –L. mollis, developed and selected from the BC1F5 progeny between wheat cv. 7182 and octoploid Tritileymus M47 (2n = 8x = 56, AABBDDNsNs), was characterized by morphological and cytogenetic identification, analysis of functional molecular markers, genomic in situ hybridization (GISH), sequential fluorescence in situ hybridization (FISH)—genomic in situ hybridization (GISH) and disease resistance evaluation. Cytological observations suggested that M11003-3-1-15-8 contained 42 chromosomes and formed 21 bivalents at meiotic metaphase I. The GISH investigations showed that line contained 40 wheat chromosomes and a pair of L. mollis chromosomes. EST-STS multiple loci markers and PLUG (PCR-based Landmark Unique Gene) markers confirmed that the introduced L. mollis chromosomes belonged to homoeologous group 7, it was designated as Lm#7Ns. While nulli-tetrasomic and sequential FISH-GISH analysis using the oligonucleotide Oligo-pSc119.2 and Oligo-pTa535 as probes revealed that the wheat 7D chromosomes were absent in M11003-3-1-15-8. Therefore, it was deduced that M11003-3-1-15-8 was a wheat–L. mollis Lm#7Ns (7D) disomic substitution line. Field disease resistance demonstrated that the introduced L. mollis chromosomes Lm#7Ns were responsible for the stripe rust resistance at the adult stage. Moreover, M11003-3-1-15-8 had a superior numbers of florets. The novel disomic substitution line M11003-3-1-15-8, could be exploited as an important genetic material in wheat resistance breeding programs and genetic resources.