Fontanesi et al. BMC Genomics 2010, 11:639 http://www.biomedcentral.com/1471-2164/11/639 Page 7 of 15 Table 3 Validated goat CNVRs using semiquantitative fluorescent multiplex-PCR (SQF-PCR) Chromosome BTA coordinates (Btau_4.0) Gain/Loss in aCGH analyses Gain/Loss in SQF-PCR analyses Target gene symbol (Ensembl entry no.) GIMAP1 (ENSBTAG00000001198) (ENSBTAG00000027170) ASIP (ENSBTAG00000034077) AHCY (ENSBTAG00000018101) NR3C2 (ENSBTAG00000027182) Gain Gain Gain Gain Gain Gain Gain/Loss Gain Gain Gain CNVR no. 25 61-62 761 761 90 4 10 13 13 17 117225479-117366050 23334056-23640961 23739149-24536154 64082600-641571862 64082600-641571862 10532314-11608092 1 Reported in [22]. 2 Coordinates of the BTA13 of the UMD 2.0 Bos taurus genome assembly. significant hits (E-value < 4E-233, identity > 89%) were obtained, and the hits included only sequences assembled in the BTA10 encompassing the goat CNVRs no. 61 and 62 or included in unassigned scaffolds (data not shown). These regions of BTA10 have been already reported to contain CNVs in different cattle breeds [51,52] (Addi- tional file 4). Analyzing by SQF-PCR goats other than those used for aCGH, we identified that this complex region (CNVRs no. 61 and 62) is affected by both gain and loss of DNA copies that did not occur specifically in the evaluated goat breeds (Additional file 7). Therefore, even if these CNVRs were classified as gain (Figure 3B) (Additional file 2), they should be considered together as gain/loss CNVRs. CVNR no. 90 on BTA17 (the largest we identified) includes the nuclear receptor subfamily 3, group C, member 2 (NR3C2) gene (ENSBTAG00000027182). PCR primers were tested on additional animals as reported for the previous two CNVRs. Only and all Girgentana goats showed gain of DNA copies in this region (Figure 3C), including the three Girgentana goats used for aCGH analyses (Addi- tional file 7), even if the signal for one of them did not trespass the averaged log2 ratio threshold of 0.175. The NR3C2 gene encodes the mineralocorticoid receptor, which mediates aldosterone actions on salt and water balance within distal nephron cells, with crucial effects on blood pressure and potassium homeostasis. The pro- tein acts as a ligand-dependent transcription factor that Figure 3 aCGH and SQF-PCR results for CNVRs nos. 25 (A), 61-62 (B), and 90 (C). For each CNVR, results were reported for three goats indicated at the top in the correspondence of the related charts and images (C1 = Camosciata delle Alpi, animal no. 1; G1 = Girgentana, animal no. 1; G2 = Girgentana, animal no. 2; MG1 = Murciano-Granadina, animal no. 1; S1 = Saanen, animal no. 1; S3 = Saanen, animal no. 3). For the aCGH experiment, images have been reported for i) log2 ratio plot of original data, ii) log2 ratio plot of summary data (pointwise averaging of all computed profiles), iii) heatmap of log2 ratios for original, smoothed/segmented, and summary data, and iv) maps of gains/losses for smoothed/ segmented and summary data (gain is indicated in orange, loss is indicated in green). Red arrows indicate regions of copy gain. Smoothed/ segmented data were obtained with several algorithms (Lowess, Wavelet, Quantreg, ruavg, CBS, CGHseg, BioHMM, cghFLasso, GLAD, and FASeg) averaged in the summary data [77]. Semiquantitative fluorescent multiplex PCR (SQF-PCR) electropherograms for fragments of three CNVRs are superimposed on that of the reference Camosciata delle Alpi goat after normalization against the control MC1R amplicon. Results obtained using DGAT1 for normalization are overlapping with those obtained with MC1R and for this reason are not reported.