Congenic mapping of a blood pressure QTL on Chromosome 16 of Dahl rats Myriam Moujahidine, Julie Dutil, Pavel Hamet, Alan Y. Deng Research Centre, Centre Hospitalier de l’Universite de Montreal (CHUM), 7-132 Pavillon Jeanne Mance, 3840, rue St. Urbain, Montreal, Quebec, H2W 1T8, Canada Received: 18 October 2001 / Accepted: 27 November 2001 Abstract. A Chromosome (Chr) 16 segment of the Dahl salt- sensitive (S) rat was shown by linkage to contain a blood pressure (BP) quantitative trait locus (QTL). To verify and further narrow down the region harboring the QTL, we made two congenic strains by replacing two segments of the S rats with the homologous segments of the Lewis (LEW) rats. The construction of these con- genic strains was facilitated by a genome-wide marker screening. The two congenic strains contained a segment in common, and BPs of both were significantly lower than that of the S strain. Consequently, a BP QTL could be localized to the overlapping region of about 49.4 centiRay (cR) including the telomere on a radiation hybrid map. Heart weights, left and right ventricular weights, kidney weights, and aortic weights over length were all significantly decreased in the congenic strains compared with the S strain. Thus, there appeared to exist an association between the effects of the QTL on BP and on cardiac, renal, and vascular hypertrophy. The S rat is an animal model that has been intensely utilized for mapping QTL (singular and plural) for BP (Deng 1998). In our previous genetic studies, a suggestive linkage was detected be- tween a region on Chr 16 and BP (Garrett et at. 1998). This was consistent with the work of other investigators using another strain of hypertensive rats (Schork et al. 1995). However, linkage alone was not sufficient to establish the authenticity of a QTL. For example, a QTL was initially localized by linkage to a segment on Chr 17 (Deng et al. 1994), but the region where the QTL most likely resided turned out not to harbor one by the use of a congenic strain (Garrett et al. 1998). Because the construction of a congenic strain involves physically moving a section of a chromosome, it provides more definitive and stringent evidence that the chromo- some region in question indeed contains a BP QTL. To prove and, if it exists, to fine map the QTL on Chr 16, we constructed two congenic strains. This report provides physical evidence for the involvement of a Chr 16 segment in BP regulation. Materials and methods Animals. The SS/Jr rat used for making congenic strains was obtained from J. Rapp and is customarily designated as S. LEW/CrlBR (LEW) rats were purchased from Charles Rivers (La Salle, Canada). These strains were maintained in our facility and monitored as reported previously (Dutil and Deng 2001). Protocols for handling as well as maintaining animals were approved by our institutional animal committee. All procedures for the experiment were in accordance with the guidelines of local, provincial, and federal regulations. Breeding scheme for generating congenic strains. The breeding pro- cedure and screening protocol were essentially the same as reported pre- viously (Deng et al. 2001). In brief, rats of the S and LEW strains were first bred to produce F 1 rats, which were backcrossed to S rats to produce the first backcross generation (BC1). BC1 rats were genotyped for 92 markers approximately evenly spaced throughout the rat genome with an average spacing of about 18 cM (Table 1). The BC1 rat, which was heterozygous SL for a Chr 16 region but possessed the maximum SS homozygosity for the rest of the irrelevant genome, was selected and designated as the ideal breeder for a subsequent breeding with an S rat to produce BC2. BC2 rats were genotyped and screened exactly as for the BC1 rats to derive an ideal breeder BC2. This process continued until BC5. At this point, it was found that only the markers delineating a Chr 16 region of interest (Fig. 1) were heterozygous SL, whereas the markers for the rest of the genome including those flanking the Chr 16 region of interest were homozygous SS. To establish a congenic strain, a BC5 rat was bred to an S rat to duplicate the segment of interest. Subsequently, a female and a male rat were sister–brother crossed to finally generate rats homozygous LL for the region of interest, but homozygous SS for the rest of Chr 16 and the rest of the genome. Two congenic strains produced are S.LEW-D16Uia2/ D16Rat12 and S.LEW-D16Mit2/D16Rat12, and are abbreviated as S.L1 and S.L2 respectively. The chromosome regions homozygous LL in each strain are shown as solid bars in Fig. 1. All the markers in the region were genotyped for both congenic strains. BP measurements. The BP study for the rat is essentially the same as described previously (Dutil and Deng 2001). In brief, the mating pairs of the S and congenic strains to be studied were bred simultaneously. Male rats were weaned at 21 days of age, maintained on a low-salt diet (0.2% NaCl, Harlan Teklad 7034), and then fed a high-salt diet (2% NaCl, Harlan Teklad 94217) starting from 35 days of age until the end of the experiment. The implantation of telemetry probes, the age of animals, and postoperative care of animals are the same as described previously (Dutil and Deng 2001; Deng et al. 2001). Tissue extraction. Rats were sacrificed by decapitation at 14 weeks of age. The organs of interest were removed, cleaned from surrounding ad- ventitial connective tissues and fat, blotted to remove excess blood, and weighed immediately. To minimize any potential inconsistency in the col- lection, one person was designated for harvesting one particular organ and also for the subsequent dissections into sub-portions of interest. The whole heart was then dissected into the left ventricle plus the septum and into the right ventricle. The weight of each section was then recorded and corrected for the body weight of the respective animal. DNA extraction and genotyping. DNA for each rat was extracted by tail biopsy using a Qiagen Genome kit, and the genotype of each rat was determined by PCR based on the methods previously published (Deng et al. 2001). Radiation hybrid mapping. A rat/hamster (RH) panel of 96 radiation hybrids was purchased from Research Genetics (Huntville, Ala.; http:// www.resgen.com/). For chromosome mapping, each marker was geno- typed with RH by PCR according to a previously published protocol (Deng Correspondence to: A. Deng; E-mail: alan.deng@umontreal.ca Mammalian Genome 13, 153–156 (2002). DOI: 10.1007/s00335-001-2138-3 © Springer-Verlag New York Inc. 2002 Incorporating Mouse Genome