Abstract
Genomic in situ hybridization (GISH) has been widely used to detect rye (Secale cereale L.) chromosomes in wheat (Triticum aestivum L.) introgression lines. The routine procedure of GISH using genomic DNA of rye as a probe is time-consuming and labor-intensive because of the preparation and labeling of genomic DNA of rye and denaturing of chromosomes and probes. In this study, new oligonucleotide probes Oligo-1162, Oligo-pSc200 and Oligo-pSc250 were developed. The three new probes can be used for non-denaturing fluorescence in situ hybridization (ND-FISH) assays and replace genomic DNA of rye as a probe to discriminate rye chromosomes in wheat backgrounds. In addition, previously developed oligonucleotide probes Oligo-pSc119.2-1, Oligo-pSc119.2-2, Oligo-pTa535-1, Oligo-pTa535-2, Oligo-pTa71-2, Oligo-pAWRC.1 and Oligo-CCS1 can also be used for ND-FISH of wheat and rye. These probes have provided an easier, faster and more cost-effective method for the FISH analysis of wheat and hybrids derived from wheat × rye.
Highlights
Rye (S. cereale L.) has been used in wheat (T. aestivum L.) breeding programs as a source of genes for resistance to abiotic stress, diseases and insects[1,2,3,4,5]
Genomic DNAs as probe, we report here the development and application of three oligonucleotide probes that can be used for non-denaturing fluorescence in situ hybridization (ND-Fluorescence in situ hybridization (FISH)) assays and eliminate the requirement of Genomic in situ hybridization (GISH) to discriminate rye from wheat chromosomes
Oligo-1162 produced hybridization signals on all 14 rye chromosomes in both octoploid triticales MA and MK, but no strong signals were observed on the 42 wheat chromosomes (Fig. 1a,b)
Summary
Rye (S. cereale L.) has been used in wheat (T. aestivum L.) breeding programs as a source of genes for resistance to abiotic stress, diseases and insects[1,2,3,4,5]. Synthetic oligonucleotides carrying a fluorescent label were developed for FISH analysis of wheat and rye chromosomes[8,9,13,14], making the process easier and more economical[14]. This has led to the development of an even simpler and more efficient technique termed non-denaturing FISH (ND-FISH) analysis[15,16]. Some oligonucleotide probes that were developed previously[14] have been tested for ND-FISH analysis
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