INVESTIGADORES
LIJAVETZKY diego Claudio
artículos
Título:
Construction and characterization of a bacterial artificial chromosome (BAC) library for the A genome of wheat
Autor/es:
LIJAVETZKY, D; MUZZI, G; WICKER, T; KELLER, B; WING, R; DUBCOVSKY, J
Revista:
GENOME
Referencias:
Año: 1999 vol. 42 p. 1176 - 1182
ISSN:
0831-2796
Resumen:
A genomic bacterial artificial chromosome (BAC) library of the A genome
of wheat has been constructed. Triticum monococcum accession DV92 was
selected for this purpose because it is a cultivated diploid wheat and
one of the parental lines used in the construction of a saturated
genetic map. Leaves from this accession were used to isolate
high-molecular-weight DNA from nuclei. This DNA was partially digested
with restriction enzyme Hind III, subjected to double size selection,
electroeluted and cloned into the pINDIGO451 BAC vector. The library
consists of 276,480 clones with an average insert size of 115 kb.
Excluding the 1.33% of empty clones and 0.14% of clones with
chloroplast DNA, the coverage of this library is 5.6 genome
equivalents. With this genome coverage the probability of having any
DNA sequence represented in this library is higher than 99.6%. Clones
were sorted in 720,384-well plates and blotted onto 15 high-density
filters. High-density filters were screened with several single or
low-copy clones and five positive BAC clones were selected for further
analysis. Since most of the T. monococcum BAC ends included repetitive
sequences, a modification was introduced into the classical
end-isolation procedure to select low copy sequences for chromosome
walking.