Diversity of a major repetitive DNA sequence in diploid and polyploid Triticeae

Abstract

About 90 members of a major tandemly repeated DNA sequence family originally described in rye as pSc119.2 have been isolated from 11 diploid and polyploid Triticeae species using primers from along the length of the sequence for PCR amplification. Alignment and similarity analysis showed that the 120-bp repeat unit family is diverse with single nucleotide changes and few insertions and deletions occurring throughout the sequence, with no characteristic genome or species-specific variants having developed during evolution of the extant genomes. Fluorescent in situ hybridization showed that each of the large blocks of the repeat at chromosomal sites harboured many variants of the 120-bp repeat. There were substantial copy number differences between genomes, with abundant sub-terminal sites in rye, interstitial sites in the B genome of wheat, and relatively few sites in the A and D genome. We conclude that sequence homogenization events have not been operative in this repeat and that the common ancestor of the Triticeae tribe had multiple sequences of the 120-bp repeat with a range of variation not unlike that seen within and between species today. This diversity has been maintained when sites are moved within the genome and in all species since their divergence within the Triticeae.

Detection of various U and M chromosomes in wheat-Aegilops biuncialis hybrids and derivatives using fluorescence in situ hybridisation and molecular markers

“…The slides were stored at -20°C. The repetitive DNA probes used for FISH were as follows: pSc119.2, a 120 bp highly repeated sequence amplified from rye genomic DNA and labelled with biotin-11-dUTP or digoxigenin-16-dUTP using PCR according to Contento et al (2005), Afa family (Nagaki et al 1995), a subclone of the pAs1 tandem repetitive sequences, labelled with biotin-11-dUTP or digoxigenin-16-dUTP using PCR, and pTa71 (Gerlach & Bedbrook 1979), labelled simultaneously with 50% digoxigenin-16-dUTP and 50% biotin-11-dUTP by nick translation. The (GAA) n microsatellite probe was amplified from Ae.…”

Introgression of chromosome segments from multiple alien species in wheat breeding lines with wheat streak mosaic virus resistance

“…intermedium, normal wheats, and wheats containing the T1BL*1RS translocation using proteolytic digestions (see Schwarzacher and Heslop-Harrison, 2000). Probes (described in the studies by Forsström et al, 2002;Contento et al, 2005;Patokar et al, 2015) included the rDNA sequences pTa71 (9 kb complete repeat unit of 25S-5.8S-18S rDNA of T. aestivum) and pTa794 (410 bp fragment of 5S rDNA of T. aestivum), and the repetitive DNA sequences pSc119.2 (or CS13, 120 bp tandem repeat isolated from S. cereale) and dpTa1 (or Afa, 340 bp tandem repeat from T. aestivum). Small insert clones were amplified by PCR using M13 primers.…”

Production and cytomolecular identification of new wheat-perennial rye (Secale cereanum) disomic addition lines with yellow rust resistance (6R) and increased arabinoxylan and protein content (1R, 4R, 6R)

“…Total genomic DNA of Kriszta rye was labelled with digoxigenin-11-dUTP using nick translation mix (Roche Diagnostics, Mannheim, Germany) and used as a probe for GISH. The FISH probe pSc119.2, a 120 bp highly repeated sequence was amplified from rye genomic DNA and labelled with biotin-11-dUTP using PCR according to Contento et al (2005). To shorten the time-consuming screening of the large amount of wheat-perennial rye progeny plants, FISH (using pSc119.2 probe) and GISH were carried out simultaneously following the protocol described by Molnár-Láng et al (2010), with minor modifications.…”