Abstract
The wild species, Thinopyrum intermedium. (Genome StStJSJSJJ), serves as a valuable germplasm resource providing novel genes for wheat improvement. In the current study, non-denaturing fluorescence in situ hybridization (ND-FISH) with multiple probes and comparative molecular markers were applied to characterize two wheat-Th. intermedium chromosome additions. Sequential ND-FISH with new labeled Th. intermedium specific oligo-probes were used to precisely determine the chromosomal constitution of Th. intermedium, wheat—Th. intermedium partial amphiploids and addition lines Hy36 and Hy37. The ND-FISH results showed that the added JS-St translocated chromosomes in Hy36 had minor Oligo-5S ribosomal DNA (rDNA) signals at the short arm, while a pair of J-St chromosomes in Hy37 had major Oligo-pTa71 and minor Oligo-5S rDNA signals. The 90K SNP array and PCR-based molecular markers that mapped on wheat linkage group 5 and 3 facilitated the identification of Thinopyrum chromosome introgressions in the addition lines, and confirmed that added chromosomes in Hy36 and Hy37 were 5JSS.3StS and 5JS.3StS, respectively. Complete coding sequences at the paralogous puroindoline-a (Pina) loci from Th. intermedium were cloned and localized on the short arm of chromosome 5JS of Hy36. Line Hy36 showed a reduction in the hardness index, which suggested that Th. intermedium-specific Pina gene sequences may be associated with the softness trait in wheat background. The molecular cytogenetic identification of novel wheat—Th. intermedium derivatives indicated that the frequent chromosome rearrangement occurred in the progenies of wheat-Thinopyrum hybridization. The new wheat-Thinopyrum derived lines may increase the genetic diversity for wheat breeding.
Highlights
Based on the comparison of current fluorescence in situ hybridization (FISH) patterns of Zhong 5 to Chinese Spring (CS) by the probes Oligo-pSc119.2 and Oligo-pTa535 [25], we found that 39 of the 42 wheat chromosomes from Zhong 5 had normal FISH patterns, except that a strong Oligo-pSc119.2 signal appeared on the end of the chromosome 1AL, and a clear Oligo-pSc119.2 signal on chromosome 2AS, as well as a distinct Oligo-pTa535 signal added on chromosome 3AS
The non-denaturing fluorescence in situ hybridization (ND-FISH) based chromosome painting technique with the labelled oligonucleotide is widely applied, due to obvious advantages over conventional FISH procedures, such as easy probes synthesis, flexible design, relatively low cost, and high-throughput [25,30,36,37,38,39]. This technique has been successfully used for distinguishing different genomes, such as Oligo-1162 for Secale cereale [30]
All 14 JS -chromosomes were clearly visualized by ND-FISH with
Summary
The great geographical and climatic diversity of the native distribution has resulted in high variability and accumulated genetic rearrangement among Th. intermedium subspecies [2]. The relative ease of crossability between wheat and Th. intermedium has led to studies involving the transfer of valuable genetic traits from. Th. intermedium to wheat [2,3]. Agronomy 2019, 9, 18 been developed [4,5,6,7,8], and a number of wheat-Th. intermedium chromosome addition, substitution, and translocation lines were produced for the localization of novel gene(s) for disease resistances and agronomic traits in diversified wheat background [9,10,11,12].
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