Isolation and characterization of X chromosome-derived DNA sequences from a dioecious plant Melandrium album

• Published in: Chromosome research Vol. 5; no. 1; pp. 57 - 65
• Main Authors: Buzek, J, Koutnikova, H, Houben, A, Riha, K, Janousek, B, Siroky, J, Grant, S, Vyskot, B
• Format: Journal Article
• Language: English
• Published: Netherlands Springer Nature B.V 01.02.1997
• Subjects: Base Composition; Base Sequence; Blotting, Northern; Blotting, Southern; Chromosome Banding; chromosome mapping; Cloning, Molecular; Deoxyribonucleic acid; DNA; DNA Methylation; DNA Probes; DNA, Plant - genetics; DNA, Plant - isolation & purification; Genes; Genetics; Heterochromatin - ultrastructure; In Situ Hybridization, Fluorescence; Meiosis - genetics; Melandrium; Mitosis - genetics; Molecular Sequence Data; nucleotide sequences; Plants - genetics; Polymerase Chain Reaction; Polymorphism, Genetic; Repetitive Sequences, Nucleic Acid; RNA, Plant - genetics; RNA, Plant - isolation & purification; Sequence Analysis, DNA; Sex Chromatin; Silene; Species Specificity; X Chromosome - genetics; X Chromosome - ultrastructure; Y Chromosome - genetics; Y Chromosome - ultrastructure
• ISSN: 0967-3849; 1573-6849
• DOI: 10.1023/a:1011693603279

A number of X chromosome DNA sequences have been isolated from a dioecious plant, Melandrium album (syn. Silene latifolia), using chromosome microdissection followed by degenerate oligonucleotideprimed polymerase chain reaction (DOP-PCR) amplification. Six DNA clones were selected and further characterized by DNA/DNA hybridization techniques to check their copy numbers, sex-specific methylation patterns, species specificity and positions on chromosomes. These clones were moderately to highly repetitive (approximately 10(3)-10(5) copies per haploid genome) and none of them gave a positive signal on Northern blots. One of the clones yielded a sex-specific methylation pattern: its abundant non-methylated CCGG island was found only in males. All the clones also hybridized to two closely related dioecious Melandrium species (M. rubrum and M. dicline). Nucleotide sequences of two X-derived clones showed a number of internal short direct repeats; one of them strikingly resembled a plant conservative telomere sequence (TTTAGGG). None of the clones hybridized to the X chromosome only, but all were localized at the telomeric heterochromatic regions (DAPI C-bands) of both arms of a vast majority of M. album chromosomes using the fluorescence in situ hybridization (FISH) technique. However, the non-homologous arm of the Y chromosome (contrary to the arm homologous to the X chromosome, possessing the pseudoautosomal region) showed neither a DAPI C-banding-stained heterochromatin nor a FISH signal with any of the DNA probes tested, thus indicating its evolutionary diversification.