Identification of Lens culinaris ssp. culinaris chromosomes by physical mapping of repetitive DNA sequences.

We describe the characterisation and the chromosomal localisation of two repeated DNA sequences, named pLc30 (466 bp long, 64% AT residues) and pLc7 (408 bp long, 61% AT residues), isolated from lentil (Lens culinaris ssp. culinaris) genomic DNA. The pLc30 family is characterised by four internal repeats organised in a head-to-tail orientation, whereas the pLc7 contains many short direct subrepeats. The two families do not share significant sequence similarity. The distribution of these repetitive sequences in different Lens species and in other legumes was investigated. pLc30 is present in all Lens species investigated but absent from other genera examined. In contrast, pLc7 is present also in the genome of other legumes. As determined by FISH, the pLc30 sequence hybridises on six out of seven lentil chromosome pairs, while pLc7 hybridises on one only. The distribution of the nine different hybridisation sites of pLc30 allows the discrimination of all seven chromosome pairs and the construction of a karyotype of L. culinaris ssp. culinaris. Additionally, the combination of simultaneous and successive FISH with pLc7, 5S rRNA, 18S-5.8S-25S rRNA genes, and a telomeric sequence allowed the assembly of a physical map based on lentil karyotype.

A family of dispersed repeats in the genome of Vicia faba: structure, chromosomal organization, redundancy modulation, and evolution.

A family of repeated DNA sequences of about 1200 bp in length and bordered by well-conserved, 18 bp inverted repeats (VfB family) was found in the nuclear genome of Vicia faba. The structure, chromosomal organization, redundancy modulation and evolution of these sequences were investigated. They are enriched in A+T base pairs (about 40% G+C) and lack any obvious internally repeated motif. A 64%-73% nucleotide sequence identity was found when pairwise comparisons between VfB sequences were carried out (average 69%). Direct repeats were not found to flank the inverted repeats that border these DNA sequences. The results obtained by hybridizing VfB repeats to Southern blots of V. faba genomic DNA digested with EcoRI indicated that these DNA elements are interspersed in the genome. The appearance of bands in these Southern blots and comparison of the structure of the sequences that flank different VfB elements showed that these repeats might be part of other, longer repeated DNA sequences. A high degree of dispersion throughout the genome was confirmed by cytological hybridization, which showed VfB sequences to be scattered along the length of all chromosomes and to be absent or rare only at heterochromatic chromosomal regions. These sequences contribute to intraspecific alterations of genomic size. Indeed, dot-blot hybridizations proved that their redundancy, which is positively correlated with the overall amount of nuclear DNA in each accession, varies between V. faba land races (27x10(3)-230x10(3) copies per 1C DNA). Southern blot hybridization of VfB repeats to restriction endonuclease-digested genomic DNAs of V. faba, V. narbonensis, V. sativa, Phaseolus coccineus, Populus deltoides, and Triticum durum revealed nucleotide sequence homology of these DNA elements, whatever the stringency conditions, only to the DNAs of Vicia species, and to a reduced extent to the DNAs of V. narbonensis and V. sativa compared with that of V. faba. It is concluded that VfB repeats might be descended from mobile DNA elements and contribute to change genomic size and organization during evolution.