Creation Of Breeding Materials Using Special Wheat Germplasms And Its Genetic Evaluation

Bread wheat(Triticum aestivum L.) is the most widely grown cereal in the world.It is the staple food for 35%of the world's population,and is becoming increasingly important in the developing world.To meet the demands for high yielding and stress-resistant wheat cultivars,it is desirable to increase the genetic base of this crop. Many wild species of the tribe Triticeae(Poaceae) are valuable sources for resistance to diseases,salinity,drought and insect pests.Wide crosses and synthetic amphiploids have played an important role in introgressing desirable traits from related species into cultivated wheat.Dwarfing polish wheat(Triticum turgidurn ssp.polonicum,DPW) is a tertraploid species(2n = 4x = 28,AABB),and was collected from Tulufan,Xinjiang, China.It is the only one dwarfing accession of T.turgidurn ssp.polonicum,with the height of about 68 cm,which is insensitive in gibberellin acid reaction,and is characterized as dwarfing,more tillers,more length of spike,more spikelets per spike and length of glume.Psathyrostachys huashanica Keng ex,Guo(2n=2x=14,NsNs) is a perennial species in the genus Psathyrostachys Nevski,and is located only in Hua Shan, Shanxi province,China.P.huashanica has been served as an important resource to wheat improvement because of its disease resistance,resistant ability to unfavorable environment,dwarfing and tremendous genetic diversity.At present,the amount of the population is much less than the past and being at the edge of extinction.Owing to distributing limitation and importance as breeding material for germplasm storage,it was listed into the first class of national protected rare plants and imperatively protected wild species in relation to crops.In the present study,we studied the genetic character and molecular mapping of dwarfing gene of dwarfing polish wheat using SSR.By colchicine treatment of the hybrid plants between dwarfing polish wheat and Aegilops tauschii Cosson.,a new synthetic hexaploid wheat(SHW-DPW) was artificially obtained for the fist time.The relationship between SHW-DPW and Triticum petropavlovskyi Udacz.et Miguch.and the origin of T. petropavlovskyi were discussed based on the sequences of a singly-copy nuclear gene encoding plastid acetyl-CoA carboxylase(Acc-1).The effects of phKL gene on crossability and homoeologous pairing of T.aestivum×p.huashanica hybrids were comparatively analyzed.The hybrid plants between T.aestivum and P.huashanica were treated with colchicine,a new intergeneric amphiploid(PHW-SA) was obtained for the fist time,and the morphology,fertility,molecular cytogenetic characterization and disease responses of the amphiploid were reported.The results were as follows:1.Dwarfing polish wheat(DPW) is a dwarfing accession of T.turgidum ssp. polonicum from Xinjiang of China.The mean plant height of DPW is 68cm.Genetic analysis of F₂,BC₁ and BC₂ populations derived from DPW×T.turgidum ssp.polonicum (AS304) indicated that the culm length in DPW was determined by a single partially dominant gene,and which was named as Rht-dp.In order to map the height-reducing gene Rht-dp,a total of 153 SSR primer pairs covering A and B wheat chromosomes were used for polymorphism survey of DPW and AS304.Among these primers,43(28.1%) showed polymorphism between the parents.By Bulk Segregant Analysis(BSA) of dwarfing and tall pools,four pairs of primer located on chromosome arm 4BS,WMC511,Xgwm495,Xgwm 113 and Xgwm251,were found being linked to Rht-dp.The 124 F₂ individuals were then amplified with the four markers.Further molecular mapping showed that Rht-dp was linked with SSR markers in the order of WMC511-Rht-dp-Xgwm495-Xgwm113-Xgwm251 with genetic distances of 0.4cM,4.1cM,2.5cM and 10.3cM using the software Mapmaker 3.0,respectively.The molecular markers developed in this study will be useful for marker-assisted selection of alternative height-reducing genes,and to better map and clone Rht-dp gene. 2.The artificial hybridization between dwarfing polish wheat and two accessions of Ae.tauschii(AS60 and AS65) was carried out,and the F₁ hybrids were obtained successfully without using embryo rescue techniques for the first time.The crossabilities of hybrids T.turgidum ssp.polonicum(DPW)×Ae.tauschii(AS60) and T.turgidum ssp. polonicurn(DPW)×Ae.tauschii(AS65) were 1.67%and 0.60%respectively.Only the hybrids of T.turgidum ssp.polonicum(DPW)×Ae.tauschii(AS60) germinated well,and 24 F₁ hybrid plants were obtained.All the F₁ hybrid plants grew vigorously,and the morphological traits were similar to bread wheat.The F₁ plants had some obvious traits inherited from the parents and were completely sterile.Chromosome pairing in the hybrid was characterized by a large number of univalents,with an average of 20.56 and 0.22 bivalents per PMC,and no ring bivalents and multivalents were observed.Furthermore,the potential value of the F₁ hybrids between T.turgidum ssp.polonicurn and Ae.tauschii for studying wheat origin and breeding are discussed.3.By colchicine treatment of the hybrid plants between T.turgidum ssp.polonicum (DPW) from Xinjiang in China and Ae.tauschii from Middle East,a new synthetic hexaploid wheat(SHW-DPW) was artificially obtained for the fist time.The seed sets of the two F₁ plants were 8.27%and 15.31%,respectively.The morphology of the SHW-DPW plants was comparable to that of the primary F₁ plants,and the SHW-DPW plants appeared more robust.All the SHW-DPW plants were uniform in morphology and had some obvious traits inherited from T.turgidurn ssp.polonicurn(DPW) and Ae.tauschii. Especially,the glumes were very soft and the rachis intemodia was short,which was different from other synthetic hexaploid wheat.Their spikes were morphologically similar to those of T.petropavlovskyi.The SHW-DPW plants were fertile,with 58.95%selfed seed set for the euhexaploids and 45.63%for the aneuploids,respectively.The meiosis of the SHW-DPW plants was quite regular,which showed a pairing configuration of 0.43 univalents,20.77 bivalents and 0.01 trivalents.The rate of chromosome association of the aneuploids was comparatively lower than that in the euploid individuals.The potential utilization for wheat improvement and study for the origin of T.petropavlovskyi are discussed in the present paper. 4.Based on the sequences of a singly-copy nuclear gene encoding plastid acetyl-CoA carboxylase(Acc-1),a total of 47 accessions Triticum and Aegilops representing diploid, tetraploid and hexaploid were used to estimate the origin of T.petropavlovskyi and the relationship of synthetic hexaploid wheat(SHW-DPW) and T.petropavlovskyi. Phylogenetic analysis was performed using maximum likelihood(ML),and median-joining (MJ) networks.Genetic diversity of all T.petropavlovskyi and T.turgidum ssp. polonicum accessions were high owing to a number of nucleotide substitutions among the wheat species.Among the three genomes,the A and B genome Acc-1 gene of T. petropavlovskyi have the more polymorphisms than D genome.The Acc-1 gene of SHW-DPW was consistently more closely related to the parent T.polonicum and Ae. tauschii than to T.petropavlovskyi,it showed that the relationship of SHW-DPW and T. petropavlovskyi was distant.Furthermore,it suggested that the A and B genome haplotypes from Acc-1 loci show that T.petropavlovskyi shares the highest average sequence identity with T.turgidum ssp.polonicum from Xinjiang and T.aestivum,and reveals specific progenitor-descendant relationships in the MJ networks based on intron sequence.In the ML tree of intron + sy data sets,the D genome sequences of the Acc-1 genes from T. petropavlovskyi are identical to the sequences of the D genome orthologs in T.aestivum, while the relationship of T.petropavlovskyi and Ae.tauschii are most distant,which is in agreement with the result of MJ networks analysis.Findings of this study reduced the probability of an independent allopolyploidization event and a single mutation in T. aestivum in the origin of T.petropavlovskyi and indicated a greater degree of gene flow between T.aestivum and T.turgidum ssp.polonicum leading to T.petropavlovskyi.It is most likely that T.petropavlovskyi was introduced from T.turgidum ssp.polonicum to T. aestivum via a spontaneous introgression or breeding effort.5.In the natural populations of common wheat landrace Kaixianluohanmai,there was a phKL gene which promotes homoeologous pairing of wheat-alien hybrids.In the present study,the effects of phKL gene on crossability and homoeologous pairing of T.aestivum×P.huashanica hybrids were comparatively analyzed.The crossability of the hybrid between Sichuan wheat landrace Kaixianluohanmai and P.huashanica was highest in all the hybrid combinations with 3.18%.The hybrids T.aestivum(Kaixianluohanmai)×p. huashanica showed a pairing configuration of 21.70 univalents + 2.68 rod bivalents + 0.34 ring bivalents + 0.06 trivalents + 0.02 quadrivalents and 3.54 chiasma per PMC at MI. However,the chiasma in hybrids of CS,CSphlb,CSph2a and CSph2b with P.huashanica was 0.56,1.90,0.87 and 0.60 respectively.Compared with the hybrids of CS,CSphlb, CSph2a and CSph2b with P.huashanica,a significant increase in the chiasma of homoeologous pairing was observed in the hybrids of T.aestivum(Kaixianluohanrnai)×p. huashanica.The effects were shown in the increment of rod bivalents,ring bivalents and trivalents and reduction of univalents.The results indicated that phKL showed a higher effect on promoting homoeologous pairing than phl and ph2 in T.aestivum×p. huashanica.The wheat landrace Kaixianluohanmai with phKL gene is probably a more desirable material for alien gene transferring than Ph2-deficiency lines.6.Wide crosses and synthetic amphiploids have played an important role in introgressing desirable traits from related species into cultivated wheat.The hybrid plants between T.aestivum cv.J-11 and P.huashanica were treated with colchicine,a new intergeneric amphiploid(PHW-SA) was obtained for the fist time.The morphological characteristics of PHW-SA resembled the parent J-11.The PHW-SA plants have purple internode and floss in basal spikelet,which are inherited from P.huashanica parent.The somatic chromosome number varied from 2n=51 to 2n=56,with 70.59%plants having 56 chromosomes.At metaphaseâ… ,PHW-SA(2n=56) showed average of 1.15 univalents, 27.34 bivalents,0.03 trivalents and 0.02 tetravalents per cell,and complete chromosome pairing was found in 49.79%of the PMCs.Lagging chromosomes and micronuclei were observed in the aneuploid cells.A survey of diseases resistance revealed that the stripe rust resistance from P.huashanica was totally expressed and powdery mildew was suppressed in the PHW-SA.The fertility of PHW-SA was 37.10%-77.38%,with average of 59.11%. PHW-SA can be used as a donor source in wheat breeding programme.7.A new wheat-P,huashanica amphiploid,PHW-SA,was characterized by molecular cytological observation and tested for stripe rust resistance in order to evaluate the potential use of the P.huashanica for wheat improvement.The PHW-SA plants were deeply covered with floss in basal spikelet and purple internode,which is characteristic of the P.huashanica parent.Feulgen staining of the somatic metaphases revealed that the chromosome number varied from 51 to 56.At metaphaseâ… ,PHW-SA(2n=56) regularly showed averagely 1.15 univalents and 27.34(24-28) bivalents per cell,and complete chromosome pairing was found in 49.79%of the pollen mother cells(PMCs),indicating a degree of cytological stability.Giemsa-C banding showed that the P.huashanica chromosomes in PHW-SA produced strong bands only in their terminal regions.Genomic in situ hybridization(GISH) analysis suggested that PHW-SA carried 10 entire P. huashanica chromosomes,36 wheat chromosomes and 10 wheat-P,huashanica translocated chromosomes.Seeds storage proteins separated by acid polyacrylamide gel electrophoresis(A-PAGE) and sodium dodecyl sulphate-polyacrylarnide gel electrophoresis(SDS-PAGE) showed that PHW-SA expressed some of P.huashanica specific gliadin and glutenin bands,and some new bands appeared.A survey of diseases resistance revealed that the stripe rust resistance from P.huashanica was totally expressed and powdery mildew was suppressed in the PHW-SA.Random amplified polymorphic DNA(RAPD) analysis verified the presence of DNA of the parents in PHW-SA.The present study showed that the amphiploid PHW-SA could serve as novel sources for transfer of valuable genes to wheat.