Establishment Of Daypyrum Villosum 6V#4S Specific Marker Linkage Map And Cultivation Of Novel Translocation Lines Of Wheat Chromosomes

Daypyrum villosum?Dasypyrum villosum?L.?P.Candary,syn.Haynaldia villosa Schur?,the related species of wheat,contains many beneficial genes.Pm97033 is a wheat-Daypyrum villosum translocation line T6V#4S·6DL carrying the powdery mildew resistance to gene PmV of Daypyrum villosum,which has high resistance to wheat powdery mildew in broad spectrum.Moreover,its gene sequence and response type to most powdery mildew strains are different from those of gene Pm21 on6V#2S.Therefore,Pm97033 is excellent resistance source of powdery mildew with great potential application value.However,because of the low transmission rate of translocated chromosome from T6V#4S·6DL in its hybrid offspring which has not been widely used in breeding.Translocation chromosome of T6V#4S·6DL were modified in a targeted way to analyze the causes of genetic instability of T6V#4S·6DL translocation chromosome and promote its utilization in breeding.So we did the following research:?1?developed molecular markers of gene PmV and Pm21 and 6A,6D for marker-assisted selection?MAS?.?2?Both alien chromosomes of translocation lines T6V#4S·6DL and T6V#2S·6AL were introduced into the genetic background of ph1b mutant to induce gene recombination between alien chromosome arms.?3?The non-target region of 6V#4S was truncated by the method of radiation treatment of pollen of translocation line T6V#4S·6DL.?4?Two wheat alien substitution lines,6V#2?6A?and 6V#4?6D?,were crossed to obtain F₂ segregating populations.A genetic linkage map of nine PCR markers for 6VS was constructed,which was used to analyze the difference of homologous sequence conservation among wheat,barley and Daypyrum villosum.And the main research results are as follows:1.Chinese Spring group 6 nulli-tetrasomic stocks,common wheat Wan7107,translocation line T6V#2S·6AL?10SR3109?and T6V#4S·6DL?Pm97033?,D.villosum accession No.1026 and D.v#2were used as the fundamental materials to screen the target markers,and the related gene sequences and target region information of chromosomes were searched on the website:http://plants.ensembl.org/Triticum aestivum/Info/Index and https://www.ncbi.nlm.nih.gov/,Finally,a total of 12 specific molecular markers to Daypyrum villosum and wheat were developed,out of which PmV-,Pm21-,6AS-,6DS-and 6AS/6DS-are 3,5,1,2,and 1,respectively.2.Using Chinese spring CSph1b mutant as the parent material,made a cross with translocation lines T6V#2S·6AL and T6V#4S·6DL respectively,and their F₁ hybrids crossed with each other.And the progeny materials were screened and identified by chromosome-specific molecular markers and genomic in situ hybridization,and novel translocation lines,3 homozygous translocation lines6V#4S·6AL,1 heterozygous translocation line T6V#4S·6AL,and 1 recombination translocation line6V#2S#4S·6DL,were obtained in the multiple cross F₃ populations.3.Different doses of ⁶⁰Co-?radiation were used to treat the mature pollen of translocation line T6V#4S·6DL.Using molecular marker and genomic in situ hybridization to screen and identify the progeny,combined with the observation of powdery mildew resistance phenotype.Finally,the target translocation lines with truncated 6V#4S were obtained including the disease-resistant plant with gene PmV,and susceptible plant without gene PmV.4.Two wheat alien substitution lines,6V#2?6A?and 6V#4?6D?,were crossed to obtain F₁ hybrids and F₂ segregating populations.The pairing behavior of two alien chromosomes,6V#2 and 6V#4,was investigated in the pollen mother cells of the F₁ hybrid by genomic in situ hybridization?GISH?,which proved that the two alien chromosomes 6V#2 and 6V#4 can pair and exchange.The separation and exchange of nine 6V short-arm-specific molecular markers in 323 individuals of F₂ populations were investigated.A genetic linkage map of nine PCR markers for 6VS was accordingly constructed based on the exchange frequencies and compared with the physical maps of wheat and barley based on homologous sequences of the markers,which showed that conservation of sequence order compared to6V was 6H and 6B>6A>6D.5.Combined with the molecular marker-assisted selection?MAS?of wheat chromosomes 6A,6D,and 6VS,three novel types of substitution lines were selected from F₂ populations including 6V#4?6A?,6V#2?6D?,and the alien recombinants.