Semi-automatic laser beam microdissection of the Y chromosome and analysis of Y chromosome DNA in a dioecious plant, Silene latifolia.
Silene latifolia has heteromorphic sex chromosomes, the X and Y chromosomes. The Y chromosome, which is thought to carry the male determining gene, was isolated by UV laser microdissection and amplified by degenerate oligonucleotide-primed PCR. In situ chromosome suppression of the amplified Y chromosome DNA in the presence of female genomic DNA as a competitor showed that the microdissected Y chromosome DNA did not specifically hybridize to the Y chromosome, but hybridized to all chromosomes. This result suggests that the Y chromosome does not contain Y chromosome-enriched repetitive sequences. A repetitive sequence in the microdissected Y chromosome, RMY1, was isolated while screening repetitive sequences in the amplified Y chromosome. Part of the nucleotide sequence shared a similarity to that of X-43.1, which was isolated from microdissected X chromosomes. Since fluorescence in situ hybridization analysis with RMY1 demonstrated that RMY1 was localized at the ends of the chromosome, RMY1 may be a subtelomeric repetitive sequence. Regarding the sex chromosomes, RMY1 was detected at both ends of the X chromosome and at one end near the pseudoautosomal region of the Y chromosome. The different localization of RMY1 on the sex chromosomes provides a clue to the problem of how the sex chromosomes arose from autosomes.
Matsunaga
S
S
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan.
Kawano
S
S
Michimoto
T
T
Higashiyama
T
T
Nakao
S
S
Sakai
A
A
Kuroiwa
T
T
D001483:Base Sequence; D017343:Genes, Plant; D005821:Genetic Techniques / Q000295:instrumentation; D017404:In Situ Hybridization, Fluorescence; D007834:Lasers; D019684:Magnoliopsida / Q000235:genetics*; D008969:Molecular Sequence Data; D012335:RNA, Ribosomal / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D019849:Sex Determination Processes; D016615:Telomere / Q000235:genetics; D014466:Ultraviolet Rays; D014960:X Chromosome / Q000235:genetics; D014998:Y Chromosome / Q000235:genetics*
D012335:RNA, Ribosomal