Characterization of repeated sequences from microdissected B chromosomes of Crepis capillaris

• Published in: Chromosoma 1995, Vol.104 (2), p.113-120
• Main Authors: Jamilena, M, Garrido-Ramos, M, Rejon, M.R, Rejon, C.R, Parker, J.S
• Format: Article
• Language: English
• Subjects: a chromosomes; accessory chromosomes; Base Sequence; Blotting, Southern; chromosomes; Chromosomes - chemistry; Cloning, Molecular; Crepis; Crepis capillaris; dna amplification; DNA Primers; DNA, Plant - chemistry; genbank/u26693; genbank/u26694; genbank/u26695; genbank/u26696; genbank/u26829; Heterochromatin; in situ hybridization; In Situ Hybridization, Fluorescence; Micromanipulation; Molecular Sequence Data; nucleic acid hybridization; nucleotide sequences; Plants - genetics; Polymerase Chain Reaction - methods; repetitive sequences; Repetitive Sequences, Nucleic Acid; Sequence Homology, Nucleic Acid; Southern blotting
• ISSN: 0009-5915; 1432-0886
• DOI: 10.1007/BF00347693

The B chromosome of Crepis capillaris was isolated from the standard chromosomes by microdissection, and the chromosomal DNA amplified using the degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). The PCR product was cloned and a B-specific library created and characterised. Southern and in situ hybridisation analyses of the DOP-PCR product from microdissected B chromosomes confirmed that the B chromosome is composed mainly of sequences also present in the A chromosomes but lacks the main repeated DNA families located in the A-chromosomal heterochromatin. From 100 clones analysed, 12% of the generated B-chromosomal library was shown to be composed of dispersed repeats located in both the A and B chromosomes. No B-specific repeated sequence was detected. One of the most abundant repeated DNAs within the library, the family B134, was further characterised. Repeating units show a sequence similarity range from 69% to 90% and are characterised by their richness in (CA)n repeats. In situ hybridisation revealed that members of this family are dispersed throughout the A and B chromosomes but are more concentrated in the pericentromeric heterochromatin of the B, indicating that the molecular organization of B heterochromatin is different from that of the A chromosomes. Compared with the A chromosomes, the Bs contain about 20,000 copies per micron more of the B134 sequence. This indicates that B134 was amplified on the B chromosome after its origin. The B134 sequences in the B chromosomes have also diverged from those on the A chromosomes. Although the DNA composition of A and B chromosomes is similar, Bs are evolving separately from A chromosomes at the molecular level.