Physical mapping of the blue-grained gene from <Emphasis Type="Italic">Thinopyrum ponticum</Emphasis> chromosome 4Ag and development of blue-grain-related molecular markers and a FISH probe based on SLAF-seq technology

Key message

A Thinopyrum ponticum chromosome 4Ag physical map was constructed, the blue-grained gene was localized, and related specific markers and a FISH probe were developed by SLAF-seq.

Abstract

Decaploid Thinopyrum ponticum (2n?=?10x?=?70) serves as an important gene pool for wheat improvement. The wheat-Th. ponticum 4Ag (4D) disomic substitution line Blue 58, derived from a distant hybridization between Th. ponticum and common wheat (Triticum aestivum L.), bears blue coloration in the aleurone layer. To map the blue-grained gene, eight wheat-Th. ponticum 4Ag translocation lines with different chromosomal segment sizes were obtained from Blue 58 using ⁶⁰Co-γ ray irradiation and were characterized using cytogenetic and molecular marker analysis. A small-segment blue-grained wheat translocation line L13, accounting for one-fifth of 4AgL, was obtained. A physical map of chromosome 4Ag was constructed containing 573 specific-locus amplified fragment sequencing (SLAF-seq) markers, including three bins with 223 markers on 4AgS and eight bins with 350 markers on 4AgL. The blue-grained gene in three blue-grained translocation lines L5, L9, and L13, was located on bin 4AgL-6 with FL 0.75–0.89. Moreover, 89 blue-grain-related molecular markers and one fluorescence in situ hybridization (FISH) probe, pThp12.19, were identified in this bin. The newly developed translocation lines and the molecular markers and FISH probe will facilitate the application of the Th. ponticum-origin blue-grained characteristic in wheat breeding.

     

Characterisation of <Emphasis Type="Italic">Thinopyrum bessarabicum</Emphasis> chromosomes through genome-wide introgressions into wheat

Key message

Genome-wide introgressions of Thinopyrum bessarabicum into wheat resulted in 12 recombinant lines. Cytological and molecular techniques allowed mapping of 1150 SNP markers across all seven chromosomes of the J genome.

Abstract

Thinopyrum bessarabicum (2n = 2x = 14, JJ) is an important source for new genetic variation for wheat improvement due to its salinity tolerance and disease resistance. Its practical utilisation in wheat improvement can be facilitated through development of genome-wide introgressions leading to a variety of different wheat–Th . bessarabicum translocation lines. In this study, we report the generation of 12 such wheat–Th . bessarabicum recombinant lines, through two different crossing strategies, which were characterized using sequential single colour and multi-colour genomic in situ hybridization (sc-GISH and mc-GISH), multi-colour fluorescent in situ hybridization (mc-FISH) and single nucleotide polymorphic (SNP) DNA markers. We also detected 13 lines containing different Th. bessarabicum chromosome aberrations through sc-GISH. Through a combination of molecular and cytological analysis of all the 25 lines containing Th. bessarabicum recombinants and chromosome aberrations we were able to physically map 1150 SNP markers onto seven Th. bessarabicum J chromosomes which were divided into 36 segmental blocks. Comparative analysis of the physical map of Th. bessarabicum and the wheat genome showed that synteny between the two species is highly conserved at the macro-level and confirmed that Th. bessarabicum contains the 4/5 translocation also present in the A genome of wheat. These wheat–Th . bessarabicum recombinant lines and SNP markers provide a useful genetic resource for wheat improvement with the latter having a wider impact as a tool for detection of introgressions from other Thinopyrum species containing the J or a closely-related genome such as Thinopyrum intermedium (J^rJ^rJ^vsJ^vsStSt) and Thinopyrum elongatum (E^eE^e), respectively.

     

Characterization of a wheat–Thinopyrum bessarabicum (T2JS-2BS·2BL) translocation line

Thinopyrum bessarabicum (2n = 2x = 14, JJ or E^bE^b) is an important genetic resource for wheat improvement due to its salinity tolerance and disease resistance. Development of wheat–Th. bessarabicum translocation lines will facilitate its practical utilization in wheat improvement. In this study, a novel wheat–Th. bessarabicum translocation line T2JS-2BS·2BL, which carries a segment of Th. bessarabicum chromosome arm 2JS was identified and further characterized using sequential chromosome C-banding, genomic in situ hybridization (GISH), dual-color fluorescent in situ hybridization (FISH) and DNA markers. The translocation breakpoint was mapped within bin C-2BS1-0.53 of chromosome 2B through marker analysis. Compared to the Chinese Spring (CS) parent and to CS-type lines, the translocation line has more fertile spikes per plant, longer spikes, more grains per spike and higher yield per plant, which suggests that the alien segment carries yield-related genes. However, plants with the translocation are also taller, head later and have lower 1,000-kernel weight than CS or CS-type lines. By using markers specific to the barley photoperiod response gene Ppd-H1, it was determined that the late heading date was conferred by a recessive allele located on the 2JS segment. In addition, four markers specific for the translocated segment were identified, which can be used for marker-aided screening.     

Biochemical and molecular diagnostics of Thinopyrum bessarabicum chromosomes in Triticum aestivum germ plasm

Thinopyrum bessarabicum (2n=2x=14, JJ) is a self-fertile salt-tolerant grass species, and its hybridization with Triticum aestivum to achieve the transfer of this attributes has been promoted. For the detection of alien introgression, development of diagnostic markers of Th. bessarabicum chromosomes in the wheat background has emerged as an important aspect in our intergeneric hybridization program. Six proteins/isozymes-high-molecular-weight glutenins, superoxide dismutase, grain esterase, β-amylase, glutamate oxaloacetate transaminase and α-amylase —were identified as positive markers for detecting the presence of Th. bessarabicum chromosomes in the advanced backcross derivatives of T. aestivum/Th. bessarabicum//n^* T. aestivum. Fluorescent in situ hybridization further enabled the detection of complete and translocated arms of Th. bessarabicum chromosomes in the T. aestivum background. These diagnostic markers served for tentatively characterizing a distinct set of Th. bessarabicum disomic additions to wheat (2n=44) and have facilitated establishing the homoeology of these added chromosomes.     

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.     

Identification of Blue-grained Wheat Translocation Lines Using Fluorescence in Situ Hybridization

The blue-grained wheat substitution line (blue 58) originated from wild hybridization between Triticum aestivum L. and Agropyron elongatum (Host) Beauv= Elytrigia elongatum (Host) Nevski= Thinopyrum ponticum (Host) Barkworth and Dewey (2n=10x=70) was irradiated and four translocation lines were screened by fluorescence in situ hybridization from the offsprings. The results obtained include the following: (1) both the two translocation lines, 9906 and 9902, have 42 chromosomes. The length of the translocated blue-grained segment was approximately one-third of the short-arm and one-half of the long-arm of the translocated wheat chromosome in 9906 and 9902, respectively, and the blue-grained translocated segment in 9902 was located on D genome; (2) both 9915 and 9904 have 44 chromosomes. One pair of chromosomes was translocated and two chromosomes from Th. ponticum were added in 9903, while two pairs of chromosomes were translocated in 9904 by blue-grained wheat segment. The location and application of blue-grained wheat translocation lines were discussed.     

Molecular cytogenetic characterization of Thinopyrum intermedium-derived wheat germplasm specifying resistance to wheat streak mosaic virus

Thinopyrum intermedium is a promising source of resistance to wheat streak mosaic virus (WSMV), a devastating disease of wheat. Three wheat germplasm lines possessing resistance to WSMV, derived from Triticum aestivum×Th. intermedium crosses, are analyzed by C-banding and genomic in situ hybridization (GISH) to determine the amount and location of alien chromatin in the transfer lines. Line CI15092 was confirmed as a disomic substitution line in which wheat chromosome 4A was replaced by Th. intermedium chromosome 4Ai?2. The other two lines, CI17766 and A29-13-3, carry an identical Robertsonian translocation chromosome in which the complete short arm of chromosome 4Ai?2 was transferred to the long arm of wheat chromosome 4A. Fluorescence in situ hybridization (FISH) using ABD genomic DNA from wheat as a probe and S genomic DNA from Pseudoroegneria stipifolia as the blocker, and vice versa, revealed that the entire short arm of the translocation was derived from the short arm of chromosome 4Ai?2 and the breakpoint was located at the centromere. Chromosomal arm ratios (L/S) of 2.12 in CI17766 and 2.15 in A29-13-3 showed that the translocated chromosome is submetacentric. This translocated chromosome is designated as T4AL?? 4Ai?2S as suggested by Friebe et al. (1991).     

Thinopyrum 7Ai-1-derived small chromatin with Barley Yellow Dwarf Virus (BYDV) resistance gene integrated into the wheat genome with retrotransposon

Thinopyrum intermedium is a useful source of resistance genes for Barley Yellow Dwarf Virus (BYDV), one of the most damaging wheat diseases. In this study, wheat/Th. intermedium translocation lines with a BYDV resistance gene were developed using the Th. intermedium 7Ai-1 chromosome. Genomic in situ hybridization (GISH), using a Th. intermedium total genomic DNA probe, enabled detection of 7Ai-1-derived small chro-matins containing a BYDV resistance gene, which were translocated onto the end of wheat chromosomes in the lines Y95011 and Y960843. Random amplified polymorphic DNA (RAPD) analyses using 120 random 10-mer primers were conducted to compare the BYDV-resistant translocation lines with susceptible lines. Two primers amplified the DNA fragments specific to the resistant line that would be useful as molecular markers to identify 7Ai-1-derived BYDV resistance chromatin in the wheat genome. Additionally, the isolated Th. intermedium-specific retrotransposon-like sequence pTi28 can be used to identify Th. intermedium chromatin transferred to the wheat genome.     

Genomic rearrangement between wheat and Thinopyrum elongatum revealed by mapped functional molecular markers

Thinopyrum elongatum serves as an excellent gene pool for wheat improvement. Genes for resistance to many biotic and abiotic stresses have been transferred from Th. elongatum to wheat through chromosome manipulation. For breeding programs, molecular markers enable screening of a large number of genotypes for alien chromosome introgressions. The main objective of the present study was to develop and characterize EST (expressed sequence tags) and PLUG (PCR-based Landmark Unique Gene) markers that can distinguish Th. elongatum chromatin from the wheat genomes. A total of 258 mapped EST primer pairs and 46 PLUG primer pairs were tested on DNA from wheat Chinese Spring (CS) and CS-Th. elongatum addition lines. The results showed that 43 primer pairs could be effectively mapped to specific Th. elongatum chromosomes. Twenty-two of the 43 markers displayed similar homoeologous chromosome locations to hexaploid wheat. Nine markers mapped to different linkage groups between wheat and Th. elongatum, while 12 makers mapped on two or three different Th. elongatum chromosomes. A comparison of molecular marker locations indicated that Th. elongatum genome was closely related to the D genome of wheat, and chromosome rearrangements and duplication had occurred in Th. elongatum and the wheat genomes. The markers will be useful in comparative gene mapping, chromosome evolutionary analysis, and gene introgression for wheat improvement using Th. elongatum accessions as gene donors.     

High-resolution cytological mapping of the long arm of chromosome 5A in common wheat using a series of deletion lines induced by gametocidal (Gc) genes of Aegilops speltoides

Gametocidal (Gc) genes of Aegilops in the background of the wheat genome lead to breakage of wheat chromosomes. The Q gene of wheat was used as a marker to select 19 deletion lines for the long arm of chromosome 5A of common wheat, Triticum aestivum cv. Chinese Spring (CS). The extents of deleted segments were cytologically estimated by the C-banding technique. The DNAs of deletion lines were hybridized with 22 DNA probes recognizing sites on the long arm of the chromosome (5AL) to determine their physical order. Based on the breeding behavior of the deletion lines, the location of a novel gene (Pv, pollen viability) affecting the viability of the male gamete was deduced. The segment translocated from 4AL to 5AL in CS was cytologically estimated to represent 13% of the total length of 5AL. Although DNA markers were almost randomly distributed along the chromosome arm, DNA markers located around the centromere and C-banded regions were obtained only rarely. Some deletion lines were highly rearranged in chromosome structure due to the effect(s) of the Gc gene. Applications of Gc genes for manipulating wheat chromosomes are discussed.     

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.

     

Visualization of Secale cereale DNA in wheat germ plasm by fluorescent in situ hybridization

Homozygous wheat/rye (1BL/1RS or 1AS/ 1RL) translocation lines have significantly contributed to wheat production, and several other wheat/rye translocation lines show a potential promise against biotic and abiotic stresses. Detecting the presence of rye at the chromosome level is feasible by C-banding and isozyme protocols, but the diagnostic strength of genomic in situ hybridization for eventually analyzing smaller DNA introgressions has greater significance. As a first step we have applied the genomic in situ hybridization technique to detect rye chromosomes in a wheat background using germ plasm of agricultural significance. By this method rye contributions to the translocations 1BL/1RS, 1AL/1RS, 5AS/5RL and 6BS/6RL could be identified. Differential labelling has further enabled the detection of rye and Thinopyrum bessarabicum chromosomes in a trigeneric hybrid of Triticum aestivum/Th. bessarabicum//Secale cereale.     

Thinopyrum bessarabicum: biochemical and cytological markers for the detection of genetic introgression in its hybrid derivatives with Triticum aestivum L.

Summary Thinopyrum bessarabicum (2n = 2x = 14, JJ) with its unique property of salt tolerance provides a potential means for the transfer of this important and complex trait into cultivated wheat through intergeneric hybridization. To accomplish this, diagnostic markers for detecting the presence of Th. bessarabicum chromosomes in a wheat background have to be established. The C-banded karyotype of Th. bessarabicum distinctly identifies individual Th. bessarabicum chromosomes and separates them from those of Triticum aestivum. Also, seven protein/isozymes, i.e., malate dehydrogenase, high-molecular-weight glutenin, Superoxide dismutase, grain esterase, glutamate oxaloacetate transaminase, -amylase and -amylase, were identified as being positive markers specific to Th. bessarabicum; these were also expressed in the T. aestivum/Th. bessarabicum amphiploid. These diagnostic biochemical markers could be useful in detecting and establishing homoeology of Th. bessarabicum chromosomes in T. aestivum/Th. bessarabicum intergeneric hybrid derivatives.     

Introgression of a novel Thinopyrum intermedium St-chromosome-specific HMW-GS gene into wheat

Thinopyrum intermedium has been hybridized extensively with wheat (Triticum aestivum L.) and several genes for disease resistance have been introgressed to cultivated wheat. However, there are very few reports about the Th. intermedium-derived seed storage protein genes which have been transferred into a wheat background by chromosome manipulation. Our aim is to identify several wheat–Th. intermedium ssp. trichophorum derivatives, and document these lines by genomic in situ hybridization (GISH), molecular markers and seed storage protein analysis. We found that a novel Th. intermedium 1St#2 chromosome-specific high-molecular-weight glutenin subunit (HMW-GS) was transferred to the wheat–Thinopyrum derivative lines. The genomic sequence of the Thinopyrum-derived HMW-GS was characterized and designated Glu-1St#2x, since it resembled x-type glutenins in both the N-terminal domain and C-terminal domain. It is much shorter than that of reported HMW-GS genes. The Glu-1St#2x sequence was successfully expressed in Escherichia coli and resulted in the identical weight to the native protein. The GISH and newly developed chromosome Thinopyrum-specific DNA markers enabled physically location of Glu-1St#2x to the region FL0.60–1.00 on Th. intermedium 1St#2L chromosome arm. Phylogenetic analysis revealed that the Glu-1St#2x evolved earlier than other x-type HMW-GS homoeologues in modern wheat genomes. The effect of Glu-1St#2x on protein content, sodium dodecyl sulphate sedimentation value and improvement of solvent retention capacity in wheat background suggested that Th. intermedium chromosome 1St#2 may have potential for improvement of wheat end-product quality.     

Development and characterization of wheat lines carrying stem rust resistance gene Sr43 derived from Thinopyrum ponticum

Key message

Wheat lines carrying Ug99-effective stem rust resistance gene Sr43 on shortened alien chromosome segments were produced using chromosome engineering, and molecular markers linked to Sr43 were identified for marker-assisted selection.

Abstract

Stem rust resistance gene Sr43, transferred into common wheat (Triticum aestivum) from Thinopyrum ponticum, is an effective gene against stem rust Ug99 races. However, this gene has not been used in wheat breeding because it is located on a large Th. ponticum 7el₂ chromosome segment, which also harbors genes for undesirable traits. The objective of this study was to eliminate excessive Th. ponticum chromatin surrounding Sr43 to make it usable in wheat breeding. The two original translocation lines KS10-2 and KS24-1 carrying Sr43 were first analyzed using simple sequence repeat (SSR) markers and florescent genomic in situ hybridization. Six SSR markers located on wheat chromosome arm 7DL were identified to be associated with the Th. ponticum chromatin in KS10-2 and KS24-1. The results confirmed that KS24-1 is a 7DS·7el₂L Robertsonian translocation as previously reported. However, KS10-2, which was previously designated as a 7el₂S·7el₂L-7DL translocation, was identified as a 7DS-7el₂S·7el₂L translocation. To reduce the Th. ponticum chromatin carrying Sr43, a BC₂F₁ population (Chinese Spring//Chinese Spring ph1bph1b*2/KS10-2) containing ph1b-induced homoeologous recombinants was developed, tested with stem rust, and genotyped with the six SSR markers identified above. Two new wheat lines (RWG33 and RWG34) carrying Sr43 on shortened alien chromosome segments (about 17.5 and 13.7 % of the translocation chromosomes, respectively) were obtained, and two molecular markers linked to Sr43 in these lines were identified. The new wheat lines with Sr43 and the closely linked markers provide new resources for improving resistance to Ug99 and other races of stem rust in wheat.     

High-resolution cytological mapping of the long arm of chromosome 5A in common wheat using a series of deletion lines induced by gametocidal (Gc) genes of Aegilops speltoides

Gametocidal (Gc) genes of Aegilops in the background of the wheat genome lead to breakage of wheat chromosomes. The Q gene of wheat was used as a marker to select 19 deletion lines for the long arm of chromosome 5A of common wheat, Triticum aestivum cv. Chinese Spring (CS). The extents of deleted segments were cytologically estimated by the C-banding technique. The DNAs of deletion lines were hybridized with 22 DNA probes recognizing sites on the long arm of the chromosome (5AL) to determine their physical order. Based on the breeding behavior of the deletion lines, the location of a novel gene (Pv, pollen viability) affecting the viability of the male gamete was deduced. The segment translocated from 4AL to 5AL in CS was cytologically estimated to represent 13% of the total length of 5AL. Although DNA markers were almost randomly distributed along the chromosome arm, DNA markers located around the centromere and C-banded regions were obtained only rarely. Some deletion lines were highly rearranged in chromosome structure due to the effect(s) of the Gc gene. Applications of Gc genes for manipulating wheat chromosomes are discussed.     

Molecular markers show a complex mosaic pattern of wheat-Thinopyrum intermedium translocations carrying resistance to YDV

Thinopyrum intermedium translocations derived from the wheat (Triticum aestivum L.) substitution line P-29 were previously characterized by RFLP. We have further analyzed these lines and additional related germplasm with publicly available STS and SSRs. Primers which showed a polymorphism between wheat and P-29, were tested in all recombinant and nulli-tetrasomic lines confirming their position on chromosome 7D. The resulting 7D/7E chromosome maps appeared as a mosaic of wheat and Th. intermedium chromatin sections. To verify the composition of the translocation lines suggested by the RFLP-PCR map, F₂ progeny of two crosses (CS/216-1 and CS/260-1) were analyzed with molecular markers. Both populations gave an unexpectedly diverse number of recombinant individuals, suggesting that interstitial translocations occur more frequently than previously thought. This analysis also showed that there is a wide range in the number and position of the interstitial translocations within a given line such as the mosaic chromosome in recombinant line 260-1/CS-26, which has four Th. intermedium chromosome segments. Phenotypic data of the two populations suggested the presence of one gene which we have called Bdv3 to differentiate it from the previously reported orthologous gene Bdv2. Using the PCR-based molecular markers identified in this study, 5 out of 12 elite lines that showed good yields and no YDV symptoms contained Th. intermedium chromatin. Due to the multiple components involved in the YDV disease complex, combining selection for YDV resistance with the molecular markers and maps identified in this study will increase the efficiency of introgressing Th. intermedium chromatin containing YDV resistance or other beneficial traits into elite wheat germplasm.     

Molecular cytogenetic identification of a wheat–<Emphasis Type="Italic">Aegilops geniculata</Emphasis> Roth 7M<Superscript>g</Superscript> 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 7M^g 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 7M^g). 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.     

Physical mapping of the blue-grained gene(s) from Thinopyrum ponticum by GISH and FISH in a set of translocation lines with different seed colors in wheat.

The original blue-grained wheat, Blue 58, was a substitution line derived from hybridization between common wheat (Triticum aestivum L., 2n=6x=42, ABD) and tall wheatgrass (Thinopyrum ponticum Liu & Wang=Agropyron elongatum, 2n=10x=70, StStEeEbEx), in which one pair of 4D chromosomes was replaced by a pair of alien 4Ag chromosomes (unknown group 4 chromosome from A. ponticum). Blue aleurone might be a useful cytological marker in chromosome engineering and wheat breeding. Cytogenetic analysis showed that blue aleurone was controlled by chromosome 4Ag. GISH analysis proved that the 4Ag was a recombination chromosome; its centromeric and pericentromeric regions were from an E-genome chromosome, but the distal regions of its two arms were from an St-genome chromosome. On its short arm, there was a major pAs1 hybridization band, which was very close to the centromere. GISH and FISH analysis in a set of translocation lines with different seed colors revealed that the gene(s) controlling the blue pigment was located on the long arm of 4Ag. It was physically mapped to the 0.71-0.80 regions (distance measured from the centromere of 4Ag). The blue color is a consequence of dosage of this small chromosome region derived from the St genome. We speculate that the blue-grained gene(s) could activate the anthocyanin biosynthetic pathway of wheat.