Degenerate Oligonucleotide Primed–Polymerase Chain Reaction‐Based Chromosome Painting of P Genome Chromosomes in Agropyron cristatum and Wheat–A. cristatum Addition Lines

Abstract

Agropyron cristatum (L.) Gaertn. (2n = 4x = 28, PPPP), a wild relative of wheat, could provide many desirable genes for wheat improvement. Microdissection and degenerate oligonucleotide primed –polymerase chain reaction (DOP–PCR) is an effective way to isolate specific sequences. The development of specific sequences and functional markers of P genome could lay the foundation for gene mapping and cloning. In this study, chromosomes 6PS and 6PL were microdissected from wheat–A. cristatum 6PS and 6PL addition lines, and DOP–PCR products from both microdissected chromosomes were used as probes to hybridize with mitotic metaphase chromosomes of diploid A. cristatum, wheat–A. cristatum addition lines 6PS, 6PL, and 6P. The results showed that the distribution patterns on A. cristatum chromosomes using DOP–PCR products of microdissected 6PS as the probe were similar to those using DOP–PCR products of microdissected 6PL as the probe. They both distributed along all chromosomes, and chromosome 6P and other chromosomes were similar according to the hybridization pattern. There were no signals on wheat chromosomes indicating that DOP–PCR products contain P‐genome specific sequences. Actually, we obtained a P‐genome specific sequence from the DOP–PCR products of 6PS, which could be used as a fluoresence in situ hybridization (FISH) probe to identify P genome chromatin in wheat–A. cristatum introgression lines. These results could provide a basis for the study of structure and evolution of P genome chromosomes. Furthermore, this study may lead to mapping and cloning of the desirable genes on 6P chromosome.

A resource of large-scale molecular markers for monitoring Agropyron cristatum chromatin introgression in wheat background based on transcriptome sequences

“…Molecular markers provide a rapid, reliable, easy, high-throughput method for detecting the alien chromosome segments. We previously showed that repeat sequence markers can detect A. cristatum alien chromosomes in a common wheat background 18 , 21 . However, repeat sequence markers are distributed throughout a whole chromosome and cannot distinguished specific chromosome fragment loci.…”

“…Han et al . (2015) used the degenerate oligonucleotide-primed (DOP)-PCR products of microdissected 6PS as a fluorescence in situ hybridization (FISH) probe to identify P genome chromatin in wheat- A. cristatum introgression lines, but the method relied on cytological techniques 21 .…”

The Genomic Variation and Differentially Expressed Genes on the 6P Chromosomes in Wheat–Agropyron cristatum Addition Lines 5113 and II-30-5 Confer Different Desirable Traits

“…GISH was performed in root tip cells to identify the 6P addition chromosomes as described by Han et al [ 73 ]. A. cristatum genomic DNA was used as a probe, and Fukuho genomic DNA was used as a blocker to detect A. cristatum chromosomes in the wheat background.…”

Identification of P genome chromosomes in Agropyron cristatum and wheat-A. cristatum derivative lines by FISH

“…Two A. cristatum- specific probes (pAcTRT1 and pAcpCR2), which distribute on the telomeric and pericentromeric regions of A. cristatum chromosomes, were used as FISH probes. The probes were recovered from degenerate oligonucleotide primed-polymerase chain reaction (DOP-PCR) products of microdissected chromosome 6PS 11,12 . The two sequences were registered in NCBI GenBank with the accession numbers KP231286 and KX390711.…”