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Construction of chromosome-specific DNA libraries covering the whole genome of field bean (Vicia faba L.)

TitleConstruction of chromosome-specific DNA libraries covering the whole genome of field bean (Vicia faba L.)
Publication TypeArticolo su Rivista peer-reviewed
Year of Publication1996
AuthorsMacas, J., Gualberti G., Nouzová M., Samec P., Lucretti Sergio, and Doležel Jaroslav
JournalChromosome Research
Pagination531 - 539
Date Published1996///
KeywordsChromosome flow sorting, Chromosome-specific, Degenerate oligonucleotide-primed polymerase chain reaction, DNA libraries, Genome mapping, Vicia faba

Recombinant DNA libraries were constructed for seven chromosome types isolated from two translocation lines of field bean (Vicia faba L.) with reconstructed karyotypes. The chromosomes were selected so that the set of libraries covers the whole V. faba genome more than once. Individual chromosome types were highly purified by flow sorting, and their DNA was amplified by degenerate oligonucleotide-primed (DOP) polymerase chain reaction (PCR) and cloned into a plasmid vector. The choice of restriction site present in PCR primer and refinement of cloning protocol resulted in high cloning efficiency and allowed generation of libraries consisting of about 106 clones from 250 or 1000 sorted chromosomes. The insert size ranged between 50 and 2200 bp and the mean length estimated in individual libraries varied between 310 and 487 bp. Hybridization of cloned fragments with labelled genomic DNA showed that about 60% of inserts represented unique or low-copy sequences. The suitability of the libraries far genome mapping was demonstrated by isolation of clones containing microsatellite motifs.


Cited By (since 1996): 31Export Date: 26 August 2010Source: Scopus

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