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Chromosome Research

, Volume 4, Issue 7, pp 531–539 | Cite as

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

  • J. MacasEmail author
  • G. Gualberti
  • M. Nouzová
  • P. Samec
  • S. Lucretti
  • J. Doležel
Article

Abstract

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 wholeV. faba genome more than once. Individual chromosome types were highly purified by flow sorting, and their DNA was amplified by degenerate oligonucleotideprimed (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 pb. 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 for genome mapping was demonstrated by isolation of clones containing microsatellite motifs.

Key words

chromosome flow sorting degenerate oligonucleotide-primed polymerase chain reaction chromosome-specific DNA libraries genome mapping Vicia faba 

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Copyright information

© Rapid Science Publishers 1996

Authors and Affiliations

  • J. Macas
    • 1
    Email author
  • G. Gualberti
    • 1
  • M. Nouzová
    • 1
  • P. Samec
    • 1
  • S. Lucretti
    • 2
  • J. Doležel
    • 3
  1. 1.Institute of Plant Molecular BiologyČeské BudějoviceCzech Republic
  2. 2.Biotechnology and Agriculture DivisionENEA, CR CasacciaRomeItaly
  3. 3.Institute of Experimental BotanyDe Montfort University Norman Borlaug Centre for Plant Science ResearchOlomoucCzech Republic

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