Northern blot analyses using probes generated from the full-length Atp7a cDNA revealed several specific hybridization bands, all of which were more intense in RNA samples extracted from duodenal enterocytes isolated from iron-deficient rats.
FISH (fluorescent in situ hybridization) analyses using probes (CEN20, TEL12q, LSI MLL) from Vvsis (Abbott Molecular Vysis, Des Plaines, IL; Abbott Diagnostics, Maidenhead, UK) were performed according to the manufacturer's instruction.
Microarrays were designed using probes from this EST library and public genomic resources.
In situ hybridization was performed as described [55] using probes from chick EST clones of Sumo1, Pias1 and Ubc9.
We have previously shown that differential expression between samples from the same species can be estimated using probes from a closely related species [45].
Absence/presence of the transgenes was confirmed by Southern hybridization analysis [24] using probes from the PM3B (bp 1231 1956 as referred to the GenBank accession AY325736) and PMI (bp 271 810 as referred to the GenBank accession AAC74685) encoding region.
By comparing the intensity of probes in each significant region to an expected distribution generated using probes from all significant regions, we established that 15 of the 71 differentiated regions are likely CNVs (at a threshold p<0.001, see methods); this includes 7/11 regions larger than 10 kb, and all three regions larger than 30 kb.
Fluorescence in situ hybridisation (FISH) was performed on TMAs using probes from the RP11 BACs library.
The interest in PH1 prompted the cloning of AGXT cDNA [ 23, 24], using probes from the orthologous rat gene [ 25].
Levels of fecal CK19 mRNA were measured using probes from the Universal Probe Library and TaqMan Master Mix in a LightCycler thermal cycler system, according to the manufacturer's instructions (Roche Diagnostics GmbH, Mannheim, Germany).
