Brief Gene Mapping Report Cytogenet Genome Res 109:534G (2005) DOI: 10.1159/000084221 Assignment 1 of the equine colony stimulating factor 1 receptor gene (CSF1R) to equine chromosome 14q15 → q16 (ECA14q15 → q16) by in situ hybridization and radiation hybrid panel mapping J. Beck, a B.P. Chowdhary b and B. Brenig a a Institute of Veterinary Medicine, Georg August University of Göttingen, Göttingen (Germany); b Department of Veterinary Anatomy and Public Health, College of Veterinary Medicine, Texas A&M University, College Station, TX (USA) Manuscript received 30 August 2004; accepted for publication by M. Schmid 29 October 2004. 1 To our knowledge this is the first time this gene has been mapped in horse. This work was supported by a grant of the Erxleben Research and Innovation Council to B. Brenig (ERIC-BR1959-2002-02). Corresponding author: Dr. Bertram Brenig Institute of Veterinary Medicine, University of Göttingen Groner Landstrasse 2, DE–37073 Göttingen (Germany) telephone: 0049-551-393383; fax: 0049-551-393392 e-mail: bbrenig@gwdg.de ABC Fax + 41 61 306 12 34 E-mail karger@karger.ch www.karger.com © 2005 S. Karger AG, Basel 1424–8581/05/1094–0534$22.00/0 Accessible online at: www.karger.com/cgr Rationale and significance The receptor of the macrophage colony-stimulating factor 1 (CSF1R) belongs to a subfamily of receptor tyrosine kinases (Roberts et al., 1988; Yarden and Ullrich, 1988). The CSF1R gene is in close vicinity to the PDGFR1 gene (platelet-derived growth factor receptor 1) which is located only a few hundred basepairs proximal in a head-to-tail orientation. Hampe et al. (1989) demonstrated that the human CSF1R gene consists of 21 small exons interrupted by introns ranging in size from 6.3 to less than 0.1 kb. Mutations in the CSF1R gene have been shown to be causative for a number of hereditary diseases and neoplastic transformations (Ridge et al., 1990; Boultwood et al., 1991). We have isolated and characterized the complete equine CSF1R gene. In agreement with the human ortholog, the equine CSF1R gene harbours 21 exons coding for a protein of 968 amino acids and spans a region of approximately 30 kb. Materials and methods Screening of an equine BAC library An equine BAC library (Godard et al., 1998) was screened by PCR using forward primer (5)-CATTGGGTTTGCTATCACTG-3)) and reverse primer (5)-AAGCCTTTGGCTTCTGAGT-3)) (Brenig et al., 2004). A single recom- binant clone (EBAB837F9) was isolated and completely sequenced. The BAC clone harbours the complete equine CSF1R gene including the 5) and 3) region. Fluorescence in situ hybridization (FISH) The equine BAC-clone harbouring CSF1R was used in fluorescence in situ hybridization experiments on equine metaphase spreads (prepared from peripheral lymphocytes) obtained from a normal, healthy stallion. Meta- phase preparations and hybridization were carried out essentially as de- scribed previously by Lichter et al. (1990). Probes (1 Ìg DNA) were labeled with digoxigenin-11-dUTP by nick translation using the DIG-Nick Translation Mix (Roche, Penzberg, Germa- ny). Labelled probes were hybridized with 10× excess of equine Cot-DNA and 7-amino-actinomycin D. Immunodetection was performed using digox- igenin-antibodies conjugated to Cy3. Chromosomes were counterstained with DAPI and examined with a Zeiss Axioplan 2 microscope (Carl Zeiss, Jena, Germany). The G-like banding pattern generated by DAPI staining was used for chromosome identification and for regional assignment of the hybridization signals. FISH experiments were carried out twice, using duplicate slides. Thirty-nine well-spread metaphases were examined and 27 showed signals on ECA14q15 → q16 on both chromatids of both chromo- somes (Fig. 1). Radiation hybrid (RH) mapping The following primers were used to analyze the 5000 rad horse × hamster RH panel (Chowdhary et al., 2003): CSF1R-primer: f: GAGTTTATGAAGGAGGTG and r: GTGTGAGA- GTGATGTTAG (Tm: 52.7; size: 174 bp) CGMP a-subunit: f: AGAGAAAAGAGAGTGTGG and r: GAATCTG- CATTGTAACAAGG (Tm: 52.7; size: 211 bp).