Thinopyrum ponticum(2n=10×=70),a wild relative of common wheat(Triticum aestivum L.),is considered an invaluable genetic resource for wheat improvement due to its abundance of genes conferring resistance to bioti...Thinopyrum ponticum(2n=10×=70),a wild relative of common wheat(Triticum aestivum L.),is considered an invaluable genetic resource for wheat improvement due to its abundance of genes conferring resistance to biotic and abiotic stresses.This study focused on the CH97 line,derived from the BC1F7 progeny of a cross between wheat cv.7182 and Th.ponticum.Cytological evidence showed that CH97 has 42 chromosomes,forming 21 bivalents at meiotic metaphase I,with the bivalents subsequently separating and moving to opposite poles during meiotic anaphase I.Through a combination of fluorescence in situ hybridization(FISH),genomic in situ hybridization(GISH),multicolor GISH(mc-GISH),and liquid array analysis,it was determined that CH97 comprises 40 wheat chromosomes and two alien chromosomes from the Ee genome of Th.ponticum,featuring the absence of a pair of 5D chromosomes and variations in 1B,6B,and 7B chromosomes.These findings confirm that CH97 is a stable wheat-Th.ponticum 5E(5D)alien disomic substitution line.Inoculation experiments revealed that CH97 exhibits high resistance to wheat powdery mildew and stripe rust throughout the growth period,in contrast to the highly susceptible common wheat parent 7182.Compared to 7182,CH97 displayed improvements in thousand-kernel weight and kernel length.Additionally,utilizing specific-locus amplified fragment sequencing(SLAF-seq)technology,chromosome 5E-specific molecular markers were developed and validated,achieving a 33.3% success rate,facilitating marker-assisted selection for disease resistance in wheat.Overall,the CH97 substitution line,with its resistance to diseases and improved agronomic traits,represents valuable new germplasm for wheat chromosome engineering and breeding.展开更多
Partial amphiploids created by crossing common wheat （Triticum aestivum L.） and Thinopyrum ponticum （Podp.） Barkworth ＆ D. R. Dewey are important intermediates in wheat breeding because of their resistance to maj...Partial amphiploids created by crossing common wheat （Triticum aestivum L.） and Thinopyrum ponticum （Podp.） Barkworth ＆ D. R. Dewey are important intermediates in wheat breeding because of their resistance to major wheat diseases. In this study, we examined the chromosome compositions of five Xiaoyan-series wheat-Th, ponticum partial amphiploids （Xiaoyan 68, Xiaoyan 693, Xiaoyan 784, Xiaoyan 7430, and Xiaoyan 7631） using GISH, multicolor-GISH, and multicolor-FISH. We found several chromosome changes in these lines. For example, wheat chromosomes 1B and 2B were added in Xiaoyan 68 and Xiaoyan 7430, respectively, while wheat chromosome 6B was eliminated from Xiaoyan 693 and Xiaoyan 7631. Chromosome rearrangements were also detected in these amphiploids, including an interspecific translocation involving chromosome 4D and some intergenomic translocations, such as A--B and A--D translocations, among wheat genomes. Analysis of the Th. ponticum chromosomes in the amphiploids showed that some lines shared the same alien chromosomes. We also evaluated these partial amphiploids for resistance to nine races of stem rust, including TTKSK （commonly known as Ug99）. Three lines, Xiaoyan 68, Xiaoyan 784, and Xiaoyan 7430, exhibited excellent resistance to all nine races, and could therefore be valuable sources of stem rust resistance in wheat breeding.展开更多
Loss of variety resistance to stripe rust （Puccinia striiformis Westend f. sp.tritici） is an important factor causing massive periodical epidemic of rust in wheat production. Creation and development of new races of...Loss of variety resistance to stripe rust （Puccinia striiformis Westend f. sp.tritici） is an important factor causing massive periodical epidemic of rust in wheat production. Creation and development of new races of rust pathogen have led to serious crisis of resistance loss in widely planted varieties. This has quickened the search for new resistance resources. Molecular marker could facilitate the identification of the location of novel genes. A line A-3 with high resistance （immune） to currently epidemic yellow rust races （CY29, 31, 32） was screened out in offspring of Triticum aestivura x Thinopyrum ponticum. Segregation in F2 and BC1 populations indicated that the resistance was controlled by two independent genes： one dominant and one recessive. SSR markers were employed to map the two resistant genes in the F2 and BC1 populations. A marker WMC477-167bp located on 2BS was linked to the dominant gene with genetic distance of 0.4 cM. Another marker WMC364-2os bp located on 7BS was linked to the recessive-resistant gene with genetic distance of 5.8 cM. The two genes identified in this paper might be two novel stripe rust resistant genes, which were temporarily designated as YrTpl and YrTp2, respectively. The tightly linking markers facilitate transfer of the two resistant genes into the new varieties to control epidemic of yellow rust.展开更多
Stem rust, caused by Puccinia graminis Pers.:Pers. f. sp. tritici Eriks. & E. Henn. (Pgt), is a destructive wheat disease and has the potential to cause significant yield losses (Olivera et al., 2015;Soko et al., ...Stem rust, caused by Puccinia graminis Pers.:Pers. f. sp. tritici Eriks. & E. Henn. (Pgt), is a destructive wheat disease and has the potential to cause significant yield losses (Olivera et al., 2015;Soko et al., 2018). The emergence of Pgt race TTKSK, virulent to the widely deployed stem rust resistance gene Sr31 and originally named as isolate Pgt-Ug99 (Pretorius et al., 2000).展开更多
Ug99, also designated as TFKSK, is a race of Puccinia graminis Pers.：Pers f. sp. tn＇tici Eriks. and E. Henn （Pgt） with broad virulence to wheat. It is the first known Pgt race possessing virulence to Sr31, a stem ...Ug99, also designated as TFKSK, is a race of Puccinia graminis Pers.：Pers f. sp. tn＇tici Eriks. and E. Henn （Pgt） with broad virulence to wheat. It is the first known Pgt race possessing virulence to Sr31, a stem rust resistance （Sr） gene deployed in wheat varieties world- wide （Singh et al., 2011 ）. Since the first detection of TFKSK in 1998, a total of 13 Ug99 variants have been identified in several African countries.展开更多
Fhb7 is a major gene that was transferred from Thinopyrum ponticum to chromosome 7D of wheat(Triticum aestivum)and confers resistance to both Fusarium head blight(FHB)and Fusarium crown rot(FCR).However,Fhb7 is tightl...Fhb7 is a major gene that was transferred from Thinopyrum ponticum to chromosome 7D of wheat(Triticum aestivum)and confers resistance to both Fusarium head blight(FHB)and Fusarium crown rot(FCR).However,Fhb7 is tightly linked to the PSY-E2 gene,which causes yellow flour,limiting its application in breeding.To break this linkage,marker K-PSY was developed for tagging PSY-E2 and used with Fhb7 markers to identify recombination between the two genes.Screening 21,000 BC1F2 backcross progeny(Chinese Spring ph1bph1b*2/SDAU 2028)revealed two Fhb7^(+)wheat-Tp7el_(2)L lines,Shannong 2–16and Shannong 16–1,that carry a desired truncated Fhb7^(+)translocation segment without PSY-E2.The two lines show levels of resistance to FHB and FCR similar to those of the original translocation line SDAU 2028,but have white flour.To facilitate Fhb7 use in wheat breeding,STS markers were developed and used to isolate Fhb7 on a truncated Tp7el_(2) translocation segment.Near-isogenic lines carrying the Fhb7^(+)segment were generated in the backgrounds of three commercial cultivars,and Fhb7^(+)lines showed increased FHB and FCR resistance without yield penalty.The breakage of the tight linkage between Fhb7 and PSY-E2 via homoeologous recombination provides genetic resources for improvement of wheat resistance to FHB and FCR and permit the large-scale deployment of Fhb7 in breeding using marker-assisted selection.展开更多
The wild decaploid species Thinopyrum ponticum(Podp.)Barkworth&D.R.Dewey is an important source of genes against biotic and abiotic stresses affecting wheat.The wheat–Th.ponticum partial amphiploid AUS6770 shows ...The wild decaploid species Thinopyrum ponticum(Podp.)Barkworth&D.R.Dewey is an important source of genes against biotic and abiotic stresses affecting wheat.The wheat–Th.ponticum partial amphiploid AUS6770 shows resistance to multiple diseases,including stripe rust,stem rust,and powdery mildew.Mitotic chromosomes of AUS6770 were characterized by non-denaturing-fluorescence in situ hybridization(ND-FISH),and the individual Th.ponticum chromosomes 1Ae to 7Ae were karyotypically distinguished by Oligo-FISH painting using bulked oligo pools based on wheat-barley collinear regions.A novel stripe rust resistant line A155,derived from AUS6770,was found to have 44 chromosomes,including a pair of 2Ae chromosomes and a pair of 6B-6Ae translocations.To detect plants with transfer of resistance genes from A155 to wheat chromosomes,1770 plants were developed from F2–F5 progenies of A155 crossed with the susceptible wheat cultivar MY11 and characterized with ND-FISH using multiple probes.A high frequency of transmission of chromosome 2Ae was observed,and 31 types of 2Ae chromosomal aberrations were identified using ND-FISH.Ten chromosomal bins on the 2Ae chromosome were determined from the deletion and translocation lines based on genome-based PCR markers.In combination with the evaluation of disease resistance,the gene(s)for stripe rust resistance was located on the FL0.79–1.00 of 2AeS and covers the corresponding region of 0–58.26 Mb in the reference genome of Th.elongatum.The newly identified wheat-Th.ponticum 2Ae translocation lines can be exploited as potential germplasm in wheat breeding for stripe rust resistance.展开更多
为了解十倍体长穗偃麦草5Ag染色体在不同小麦背景中的遗传稳定性及其在配子中的传递,本研究利用小麦-十倍体长穗偃麦草二体代换系DS5Ag(5D),即百农普偃5814(PY5814),与6个小麦品种(系)中国春、百农矮抗58、兰考矮早8、温麦6号、周麦16以...为了解十倍体长穗偃麦草5Ag染色体在不同小麦背景中的遗传稳定性及其在配子中的传递,本研究利用小麦-十倍体长穗偃麦草二体代换系DS5Ag(5D),即百农普偃5814(PY5814),与6个小麦品种(系)中国春、百农矮抗58、兰考矮早8、温麦6号、周麦16以及Q03073进行杂交,杂种F1自交,并分别与上述6个品种(系)进行正反交,利用分子标记技术对F2和BC1F1代进行鉴定。结果表明,PY5814与6个不同小麦品种(系)杂交获得的F1均能够正常结实;在不同组合的F2分离群体中,5Ag染色体传递率范围为51.61%~71.76%,表明5Ag染色体在不同小麦背景中的遗传率不同,传递受小麦背景的影响。在BC1F1群体中,5Ag染色体通过雌配子和雄配子的传递率范围分别为21.43%~31.73%和29.51%~39.73%,表明5Ag染色体可以通过雌、雄配子传递;在兰考矮早8和温麦6号背景下,其通过雄配子的传递率高于通过雌配子的传递率,表明其在遗传研究中适合做父本。同时利用基因组荧光原位杂交(GISH,genomic in situ hybridization)技术对部分F2材料进行了验证,其结果与分子标记结果一致。本研究还为构建适宜杂交群体规模的预测,从而在后代中有效筛选出携带5Ag染色体的目标单株提供了理论指导。展开更多
Blue-grained wheat derived from the hybrid Triticum aestivum L. X Thinopyrum ponticum (Podp.) Barkworth et D. R. Dewey (Agropyron elongatum (Host) P. Beauv., 2n=70). The molecular biological mechanism of the biosynthe...Blue-grained wheat derived from the hybrid Triticum aestivum L. X Thinopyrum ponticum (Podp.) Barkworth et D. R. Dewey (Agropyron elongatum (Host) P. Beauv., 2n=70). The molecular biological mechanism of the biosynthetic pathway of blue pigments in the blue grain remains unclear yet. Dihydroflavonol 4-reductase (DFR) is one of the key enzymes controlling flavonoid synthesis in anthocyanin biosynthetic pathway, and may directly participate in the formation of blue pigment in the aleurone layer of blue-grained wheat. Here we cloned a DFR cDNA (TaDFR) from the developing seeds of blue-grained wheat, and four DFR genomic DNAs from Th. ponticum (ThpDFR.t), blue-grained wheat (TaDFR.bg), white-grained offspring of light blue-grained wheat (TaDFR.wg) and Chinese Spring (2n=42) (TaDFR.csg), respectively. TaDFR cDNA encodes a 354 amino-acids polypeptide with high identity to DFR from Hordeum vulgare L. (94%), Oryza sativa L. (83%), Zea mays L.(84%). The result of cluster analysis showed that TaDFR cDNA nucleotide sequence has 100% identity with that of TaDFR.csg. The four DFR genomic DNAs have extraordinary high homology and each has three introns. The differences of the four DFR genomic DNAs mainly exist in introns. Southern blotting analysis showed that there are at least 3-5 DFR copies in wheat, the copy numbers in different color grain wheats are not significantly different. The hybridization band patterns were the same, but different from that of Th. ponticum. DFR in blue-grained wheat belongs to a DFR superfamily. Northern blotting analysis indicated that the DFR expressed in the developing seeds of both blue- and white-grained wheat at 15 d after flowering (DAF), the mRNA levels of DFR reached the highest at 18 DAF, then declined quickly and disappeared at 33 DAF But the expression levels in blue-grained seeds were higher than that in white grain at the same seed developing stages. DFR transcripts accumulated in young leaves, and leaf sheaths of blue- and white-grained wheat and Th ponticum, but not detected in roots from different color wheats and developing seeds of Th. ponticum. Results indicated that there may exist some regulatory gene(s) which can increase the expression of DFR in the aleurone layer of blue-grained wheat, and thus resulting in the formation of blue pigments.展开更多
基金funded by the Key R&D Program of Yangling Seed Industry Innovation,China(Ylzy-xm-02)the Young Elite Scientists Sponsorship Program by China Association for Science and Technology(2021QNRC001)。
文摘Thinopyrum ponticum(2n=10×=70),a wild relative of common wheat(Triticum aestivum L.),is considered an invaluable genetic resource for wheat improvement due to its abundance of genes conferring resistance to biotic and abiotic stresses.This study focused on the CH97 line,derived from the BC1F7 progeny of a cross between wheat cv.7182 and Th.ponticum.Cytological evidence showed that CH97 has 42 chromosomes,forming 21 bivalents at meiotic metaphase I,with the bivalents subsequently separating and moving to opposite poles during meiotic anaphase I.Through a combination of fluorescence in situ hybridization(FISH),genomic in situ hybridization(GISH),multicolor GISH(mc-GISH),and liquid array analysis,it was determined that CH97 comprises 40 wheat chromosomes and two alien chromosomes from the Ee genome of Th.ponticum,featuring the absence of a pair of 5D chromosomes and variations in 1B,6B,and 7B chromosomes.These findings confirm that CH97 is a stable wheat-Th.ponticum 5E(5D)alien disomic substitution line.Inoculation experiments revealed that CH97 exhibits high resistance to wheat powdery mildew and stripe rust throughout the growth period,in contrast to the highly susceptible common wheat parent 7182.Compared to 7182,CH97 displayed improvements in thousand-kernel weight and kernel length.Additionally,utilizing specific-locus amplified fragment sequencing(SLAF-seq)technology,chromosome 5E-specific molecular markers were developed and validated,achieving a 33.3% success rate,facilitating marker-assisted selection for disease resistance in wheat.Overall,the CH97 substitution line,with its resistance to diseases and improved agronomic traits,represents valuable new germplasm for wheat chromosome engineering and breeding.
基金supported by the grants from the National Natural Science Foundation of China(No.31171539)the National High-Tech Research and Development Program of China(No.2011AA1001)the National Key Technology R&D Program of China(No.2013BAD05B01)
文摘Partial amphiploids created by crossing common wheat （Triticum aestivum L.） and Thinopyrum ponticum （Podp.） Barkworth ＆ D. R. Dewey are important intermediates in wheat breeding because of their resistance to major wheat diseases. In this study, we examined the chromosome compositions of five Xiaoyan-series wheat-Th, ponticum partial amphiploids （Xiaoyan 68, Xiaoyan 693, Xiaoyan 784, Xiaoyan 7430, and Xiaoyan 7631） using GISH, multicolor-GISH, and multicolor-FISH. We found several chromosome changes in these lines. For example, wheat chromosomes 1B and 2B were added in Xiaoyan 68 and Xiaoyan 7430, respectively, while wheat chromosome 6B was eliminated from Xiaoyan 693 and Xiaoyan 7631. Chromosome rearrangements were also detected in these amphiploids, including an interspecific translocation involving chromosome 4D and some intergenomic translocations, such as A--B and A--D translocations, among wheat genomes. Analysis of the Th. ponticum chromosomes in the amphiploids showed that some lines shared the same alien chromosomes. We also evaluated these partial amphiploids for resistance to nine races of stem rust, including TTKSK （commonly known as Ug99）. Three lines, Xiaoyan 68, Xiaoyan 784, and Xiaoyan 7430, exhibited excellent resistance to all nine races, and could therefore be valuable sources of stem rust resistance in wheat breeding.
文摘Loss of variety resistance to stripe rust （Puccinia striiformis Westend f. sp.tritici） is an important factor causing massive periodical epidemic of rust in wheat production. Creation and development of new races of rust pathogen have led to serious crisis of resistance loss in widely planted varieties. This has quickened the search for new resistance resources. Molecular marker could facilitate the identification of the location of novel genes. A line A-3 with high resistance （immune） to currently epidemic yellow rust races （CY29, 31, 32） was screened out in offspring of Triticum aestivura x Thinopyrum ponticum. Segregation in F2 and BC1 populations indicated that the resistance was controlled by two independent genes： one dominant and one recessive. SSR markers were employed to map the two resistant genes in the F2 and BC1 populations. A marker WMC477-167bp located on 2BS was linked to the dominant gene with genetic distance of 0.4 cM. Another marker WMC364-2os bp located on 7BS was linked to the recessive-resistant gene with genetic distance of 5.8 cM. The two genes identified in this paper might be two novel stripe rust resistant genes, which were temporarily designated as YrTpl and YrTp2, respectively. The tightly linking markers facilitate transfer of the two resistant genes into the new varieties to control epidemic of yellow rust.
基金supported by the Natural Science Foundation of China (No. 31571644)the National Key Basic Research Program (No. 2015CB150106)the Ministry of Science and Technology of China (2014DFA31540)
文摘Stem rust, caused by Puccinia graminis Pers.:Pers. f. sp. tritici Eriks. & E. Henn. (Pgt), is a destructive wheat disease and has the potential to cause significant yield losses (Olivera et al., 2015;Soko et al., 2018). The emergence of Pgt race TTKSK, virulent to the widely deployed stem rust resistance gene Sr31 and originally named as isolate Pgt-Ug99 (Pretorius et al., 2000).
基金supported by the Ministry of Science and Technology of China (No. 2014DFA31540)Chinese Academy of Sciences (No. SAJC201305)the Bill & Melinda Gates Foundation to Cornell University for the Borlaug Global Rust Initiative (BGRI) Durable Rust Resistance in Wheat (DRRW) Project
文摘Ug99, also designated as TFKSK, is a race of Puccinia graminis Pers.：Pers f. sp. tn＇tici Eriks. and E. Henn （Pgt） with broad virulence to wheat. It is the first known Pgt race possessing virulence to Sr31, a stem rust resistance （Sr） gene deployed in wheat varieties world- wide （Singh et al., 2011 ）. Since the first detection of TFKSK in 1998, a total of 13 Ug99 variants have been identified in several African countries.
基金supported by the National Natural Science Foundation of China(32030081,31871610)the Agricultural Variety Improvement Project of Shandong Province(2019LZGC016)the U.S.Wheat and Barley Scab Initiative。
文摘Fhb7 is a major gene that was transferred from Thinopyrum ponticum to chromosome 7D of wheat(Triticum aestivum)and confers resistance to both Fusarium head blight(FHB)and Fusarium crown rot(FCR).However,Fhb7 is tightly linked to the PSY-E2 gene,which causes yellow flour,limiting its application in breeding.To break this linkage,marker K-PSY was developed for tagging PSY-E2 and used with Fhb7 markers to identify recombination between the two genes.Screening 21,000 BC1F2 backcross progeny(Chinese Spring ph1bph1b*2/SDAU 2028)revealed two Fhb7^(+)wheat-Tp7el_(2)L lines,Shannong 2–16and Shannong 16–1,that carry a desired truncated Fhb7^(+)translocation segment without PSY-E2.The two lines show levels of resistance to FHB and FCR similar to those of the original translocation line SDAU 2028,but have white flour.To facilitate Fhb7 use in wheat breeding,STS markers were developed and used to isolate Fhb7 on a truncated Tp7el_(2) translocation segment.Near-isogenic lines carrying the Fhb7^(+)segment were generated in the backgrounds of three commercial cultivars,and Fhb7^(+)lines showed increased FHB and FCR resistance without yield penalty.The breakage of the tight linkage between Fhb7 and PSY-E2 via homoeologous recombination provides genetic resources for improvement of wheat resistance to FHB and FCR and permit the large-scale deployment of Fhb7 in breeding using marker-assisted selection.
基金supported by the National Natural Science Foundation of China (31971886)International Cooperation Program Program (2022YFH0012)of the Science and Technology Department of Sichuan.
文摘The wild decaploid species Thinopyrum ponticum(Podp.)Barkworth&D.R.Dewey is an important source of genes against biotic and abiotic stresses affecting wheat.The wheat–Th.ponticum partial amphiploid AUS6770 shows resistance to multiple diseases,including stripe rust,stem rust,and powdery mildew.Mitotic chromosomes of AUS6770 were characterized by non-denaturing-fluorescence in situ hybridization(ND-FISH),and the individual Th.ponticum chromosomes 1Ae to 7Ae were karyotypically distinguished by Oligo-FISH painting using bulked oligo pools based on wheat-barley collinear regions.A novel stripe rust resistant line A155,derived from AUS6770,was found to have 44 chromosomes,including a pair of 2Ae chromosomes and a pair of 6B-6Ae translocations.To detect plants with transfer of resistance genes from A155 to wheat chromosomes,1770 plants were developed from F2–F5 progenies of A155 crossed with the susceptible wheat cultivar MY11 and characterized with ND-FISH using multiple probes.A high frequency of transmission of chromosome 2Ae was observed,and 31 types of 2Ae chromosomal aberrations were identified using ND-FISH.Ten chromosomal bins on the 2Ae chromosome were determined from the deletion and translocation lines based on genome-based PCR markers.In combination with the evaluation of disease resistance,the gene(s)for stripe rust resistance was located on the FL0.79–1.00 of 2AeS and covers the corresponding region of 0–58.26 Mb in the reference genome of Th.elongatum.The newly identified wheat-Th.ponticum 2Ae translocation lines can be exploited as potential germplasm in wheat breeding for stripe rust resistance.
文摘为了解十倍体长穗偃麦草5Ag染色体在不同小麦背景中的遗传稳定性及其在配子中的传递,本研究利用小麦-十倍体长穗偃麦草二体代换系DS5Ag(5D),即百农普偃5814(PY5814),与6个小麦品种(系)中国春、百农矮抗58、兰考矮早8、温麦6号、周麦16以及Q03073进行杂交,杂种F1自交,并分别与上述6个品种(系)进行正反交,利用分子标记技术对F2和BC1F1代进行鉴定。结果表明,PY5814与6个不同小麦品种(系)杂交获得的F1均能够正常结实;在不同组合的F2分离群体中,5Ag染色体传递率范围为51.61%~71.76%,表明5Ag染色体在不同小麦背景中的遗传率不同,传递受小麦背景的影响。在BC1F1群体中,5Ag染色体通过雌配子和雄配子的传递率范围分别为21.43%~31.73%和29.51%~39.73%,表明5Ag染色体可以通过雌、雄配子传递;在兰考矮早8和温麦6号背景下,其通过雄配子的传递率高于通过雌配子的传递率,表明其在遗传研究中适合做父本。同时利用基因组荧光原位杂交(GISH,genomic in situ hybridization)技术对部分F2材料进行了验证,其结果与分子标记结果一致。本研究还为构建适宜杂交群体规模的预测,从而在后代中有效筛选出携带5Ag染色体的目标单株提供了理论指导。
文摘Blue-grained wheat derived from the hybrid Triticum aestivum L. X Thinopyrum ponticum (Podp.) Barkworth et D. R. Dewey (Agropyron elongatum (Host) P. Beauv., 2n=70). The molecular biological mechanism of the biosynthetic pathway of blue pigments in the blue grain remains unclear yet. Dihydroflavonol 4-reductase (DFR) is one of the key enzymes controlling flavonoid synthesis in anthocyanin biosynthetic pathway, and may directly participate in the formation of blue pigment in the aleurone layer of blue-grained wheat. Here we cloned a DFR cDNA (TaDFR) from the developing seeds of blue-grained wheat, and four DFR genomic DNAs from Th. ponticum (ThpDFR.t), blue-grained wheat (TaDFR.bg), white-grained offspring of light blue-grained wheat (TaDFR.wg) and Chinese Spring (2n=42) (TaDFR.csg), respectively. TaDFR cDNA encodes a 354 amino-acids polypeptide with high identity to DFR from Hordeum vulgare L. (94%), Oryza sativa L. (83%), Zea mays L.(84%). The result of cluster analysis showed that TaDFR cDNA nucleotide sequence has 100% identity with that of TaDFR.csg. The four DFR genomic DNAs have extraordinary high homology and each has three introns. The differences of the four DFR genomic DNAs mainly exist in introns. Southern blotting analysis showed that there are at least 3-5 DFR copies in wheat, the copy numbers in different color grain wheats are not significantly different. The hybridization band patterns were the same, but different from that of Th. ponticum. DFR in blue-grained wheat belongs to a DFR superfamily. Northern blotting analysis indicated that the DFR expressed in the developing seeds of both blue- and white-grained wheat at 15 d after flowering (DAF), the mRNA levels of DFR reached the highest at 18 DAF, then declined quickly and disappeared at 33 DAF But the expression levels in blue-grained seeds were higher than that in white grain at the same seed developing stages. DFR transcripts accumulated in young leaves, and leaf sheaths of blue- and white-grained wheat and Th ponticum, but not detected in roots from different color wheats and developing seeds of Th. ponticum. Results indicated that there may exist some regulatory gene(s) which can increase the expression of DFR in the aleurone layer of blue-grained wheat, and thus resulting in the formation of blue pigments.
