Molecular cytogenetic characterisation of the terminal heterochromatic segment of the B-chromosome of rye (Secale cereale)

The terminal heterochromatic segments of the long arms of 20 rye B-chromosomes were isolated by means of laser microdissection technology. Also the remaining portions of the long arms, along with the short arms of the same chromosomes were isolated. Each sample was used for degenerate oligonucleotide primer-polymerase chain reaction (DOP-PCR) amplification reactions. The resulting products were used as probes for chromosome in situ hybridisation experiments, and in Southern hybridisation to digests of OB and +B DNA. Competition hybridisation of these probes with OB DNA allowed the detection of B-specific sequences. The terminal heterochromatin of the rye B-chromosome contains both B-specific sequences and sequences also present on the A-chromosomes of rye. The B-specific D1100 family is the major repeat species located in the terminal heterochromatin. Primers designed to the cloned sequence (E1100) were used to search for related low copy sequences in OB DNA. The sequences of the PCR products revealed no similarities to that of the clone E1100 except for the primer sequences. The possible origin of this sequence is discussed in the context of models for the evolution of the rye B-chromosome.