BAC clones labeling was carried out with a first amplification by DOP-PCR, followed by a conventional PCR for labeling, as described previously in Garcia Angulo et al.28 (link). Three different fluorochromes were used: Texas red (Thermo Fisher Scientific, USA), fluorescein-isothiocyanate (FITC) (Enzo, USA), and diethyl-aminocoumarin (DEAC) (Vysis, USA). The chromosomes were pretreated with pepsin and fixed in formaldehyde. Finally, the chromosome preparation was dehydrated with ethanol series and air-dried before hybridization. Hybridization was done according to Portela-Bens et al.29 (link).
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