Rapid identification of Psathyrostachys huashanica Keng chromosomes in wheat background based on ND-FISH and SNP array methods

Psathyrostachys huashanica Keng(2n=2x=14,NsNs)is regarded as a valuable wild relative species for common wheat cultivar improvement because of its abundant beneficial agronomic traits.However,although the development of many wheat–P.huashanica-derived lines provides a germplasm base for the transfer of excellent traits,the lag in the identification of P.huashanica chromosomes in the wheat background has limited the study of these lines.In this study,three novel nondenaturing fluorescence in situ hybridization(ND-FISH)-positive oligo probes were developed.Among them,HS-TZ3 and HS-TZ4 could specifically hybridize with P.huashanica chromosomes,mainly in the telomere area,and HS-CHTZ5 could hybridize with the chromosomal centromere area.We sequentially constructed a P.huashanica FISH karyotype and idiogram that helped identify the homologous groups of introduced P.huashanica chromosomes.In detail,1Ns and 2Ns had opposite signals on the short and long arms,3Ns,4Ns,and 7Ns had superposed two-color signals,5Ns and 6Ns had fluorescent signals only on their short arms,and 7Ns had signals on the intercalary of the long arm.In addition,we evaluated different ways to identify alien introgression lines by using low-density single nucleotide polymorphism(SNP)arrays and recommended the SNP homozygosity rate in each chromosome as a statistical pattern.The 15K SNP array is widely applicable for addition,substitution,and translocation lines,and the 40K SNP array is the most accurate for recognizing transposed intervals between wheat and alien chromosomes.Our research provided convenient methods to distinguish the homologous group of P.huashanica chromosomes in a common wheat background based on ND-FISH and SNP arrays,which is of great significance for efficiently identifying wheat–P.huashanica-derived lines and the further application of Ns chromosomes.

Identification of genetic locus with resistance to take-all in the wheat-Psathyrostachys huashanica Keng introgression line H148

Take-all is a devastating soil-borne disease of wheat(Triticum aestivum L.).Cultivating resistant line is an important measure to control this disease.Psathyrostachys huashanica Keng is a valuable germplasm resource with high resistance to take-all.This study reported on a wheat-/R huashanica introgression line H148 with improved take-all resistance compared with its susceptible parent 7182.To elucidate the genetic mechanism of resistance in H148,the F_(2)genetic segregating population of H148×XN585 was constructed.The mixed genetic model analysis showed that the take-all resistance was controlled by two major genes with additive,dominant and epistasis effects.Bulked segregant analysis combined with wheat axiom 660K genotyping array analysis showed the polymorphic SNPs with take-all resistance from P.huashanica alien introgression were mainly distributed on the chromosome 2A.Genotyping of the F_(2)population using the KASP marker mapped a major QTL in an interval of 68.8-70.1 Mb on 2AS.Sixty-two genes were found in the target interval of the Chinese Spring reference genome sequence.According to the functional annotation of genes,two protein genes that can improve the systematic resistance of plant roots were predicted as candidate genes.The development of wheat-P.huashanica introgression line H148 and the resistant QTL mapping information are expected to provide some valuable references for the fine mapping of disease-resistance gene and development of take-all resistant varieties through molecular marker-assisted selection.

Genetic structure and differentiation of Psathyrostachys huashanica populations detected with RAPD markers

Psathyrostachys huashanica Keng is a perennial grass and belongs to genus Psathyrostachys under Triticeae.sathyrostachys is found in the center of Middle Asia and the Caucasus Mountain,while P.huashanica,a species endemic to China,is only located in Mt.Hua in the Shaanxi province,China.At present,the population of this species is decreasing,and reaching the edge of extinction.Due to the limitation in distribution and the importance as breeding material for germplasm storage,it has been considered as first class among the national protected rare plants.For this reason,the present study is significant in probing plant flora,origin and evolu-tion of Triticeae,and crop breeding.Randomly amplified polymorphic DNA(RAPD)markers were used to analyze the genetic structure and differentiation of P.huashanica popula-tions sampled in three valleys(Huangpu,Xian,and Huashan Valleys)in Mt.Hua.One hundred and twenty-two RAPD fragments were obtained in all 266 individuals with 20 prim-ers with a mean of 6.1(2-10)fragments per primer.The percentage of polymorphic loci(PPB)was 60.66%in Huangpu Valley,90.98%in Xian Valley,95.08%in Huashan Valley,and the total PPB was 95.08%,which indicated a highly genetic variability of P.huashanica.The Shannon’s Information index and GST were 0.3306 and 0.3263,respectively,indicating that there were more genetic variations within the subpopulations than those among the subpopulations.The gene flow among the subpopulations of P.huashanica(Nm=1.0322)was much less than that of the common ane-mophytes(Nm=5.24).Mean genetic distance is 0.1571(range:0.0022-0.2901).The highest value of genetic distance was found between the subpopulation(hp1)of Huangpu Valley and the highest altitude subpopulation(h8)of Huashan Valley.Correlation analysis detected significant correlation between genetic distance and vertical distance of altitude.Clustering analysis and principal coordinate analysis revealed the genetic differentiation among the populations of P.huashanica.Differentiation mainly occurred between the higher altitude subpopulations and the lower altitude sub-populations,suggesting that altitude might be the major factor that restricted the gene flow between different altitude subpopulations and resulted in differentiation of subpopulations.

小麦-华山新麦草易位系H1684细胞学鉴定及抗条锈性遗传分析

通过对华山新麦草与‘7182’杂交获得的抗条锈病新种质‘H1684’进行遗传分析,明确‘H1684’含有的抗病基因遗传特点。本研究利用基因组原位杂交技术对‘H1684’含有的外源染色体片段进行鉴定,并以‘H1684’、感病对照‘铭贤169’及其杂交后代F1、F2、F2:3和BC1群体为材料,采用10个条锈菌生理小种CYR-23、CYR25、CYR27、CYR28、CYR29、CYR30、CYR31、CYR32、CYR33和CYR34对供试群体进行苗期抗条锈性鉴定,分析抗病基因的遗传规律。GISH分析表明‘H1684’含有来自于华山新麦草的长染色体片段,为华山新麦草易位系。抗病性鉴定表明:‘H1684’在苗期对10个条锈菌生理小种均表现免疫或高抗。‘H1684’对CYR33和CYR34的抗病性都是由1对显性基因控制。华山新麦草易位系‘H1684’对中国目前流行的小麦条锈菌生理小种具有良好的抗病性,可以作为抗源在中国小麦抗条锈病育种中应用。