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The bacterial artificial chromosome (BAC) library of the narrow-leafed lupin (Lupinus angustifolius L.)

Abstract

The narrow-leafed lupin possesses valuable traits for environment-friendly agriculture and for the production of unconventional agricultural products. Despite various genetic and environmental studies, the breeding of improved cultivars has been slow due to the limited knowledge of its genomic structure. Further advances in genomics require, among other things, the availability of a genomic DNA library with large inserts. We report here on the construction of the first DNA library cloned in a BAC (bacterial artificial chromosome) vector from diploid Lupinus angustifolius L. cv. Sonet. The high molecular weight DNA used for its preparation was isolated from interphase nuclei that were purified by flow cytometry. The library comprises 55,296 clones and is ordered in 144×384-well microtitre plates. With an average insert size of 100 kb, the library represents six haploid genome equivalents. Thanks to the purification of the nuclei by flow cytometry, contamination with chloroplast DNA and mitochondrial DNA was negligible. The availability of a BAC library opens avenues for the development of a physical contig map and positional gene cloning, as well as for the analysis of the plant’s genome structure and evolution.

Abbreviations

BAC:

bacterial artificial chromosome

DAPI:

4′,6-diamidino-2-phenylindole

FISH:

fluorescence in situ hybridization

HMW:

high molecular weight

PAC:

artificial bacteriophage P1 chromosome

PFGE:

pulsed field gel electrophoresis

PRINS:

primed in situ DNA labeling

YAC:

yeast artificial chromosome

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Correspondence to Bogdan Wolko.

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Key words

  • BAC
  • Genomic DNA library
  • Lupinus angustifolius
  • Narrow-leafed lupin