Interval mapping and congenic strains for a blood pressure QTL on rat Chromosome 13

The renin locus (Ren) on rat Chromosome (Chr) 13 had previously been shown to cosegregate with blood pressure in crosses involving Dahl salt-sensitive (S) and Dahl salt-resistant (R) rats. In the present work, interval mapping of blood pressure on Chr 13 with a large F₂ (S × R), n = 233, population yielded a maximum LOD = 4.2 for linkage to blood pressure, but the quantitative trait locus (QTL) was only poorly localized to a large 35-centiMorgan (cM) segment of Chr 13. In the linkage analysis, the S-rat QTL allele (S) was associated with higher, and the R-rat QTL allele (R) with lower blood pressure, the difference between homozygotes being about 20 mm Hg. A congenic strain was made by introgressing the R-rat Ren allele into the recipient S strain. This congenic strain showed a 24 mm Hg reduction (P = 0.004) in blood pressure compared with S rats for rats fed 2% NaCl diet for 24 days; this difference was confirmed by two other independent tests. Two congenic substrains were derived from the first congenic strain with shorter R Chr 13 segments on the S background. Comparisons among these congenic strains showed that a blood pressure QTL was in the 24-cM chromosomal segment between Syt2 and D13M1Mit108. This segment does not include the renin locus, which is thus excluded from being the gene on rat Chr 13 responsible for genetic differences in blood pressure detected by linkage analysis. Received: 20 December 1996 / Accepted: 7 April 1997     

Multiple blood pressure QTL on rat Chromosome 2 defined by congenic Dahl rats

Linkage analysis previously demonstrated a blood pressure quantitative trait locus (QTL) on rat Chromosome 2 (Chr 2) in crosses utilizing Dahl salt-sensitive (S) rats. The present work dissects this QTL by using congenic strains in which segments of Chr 2 from Wistar Kyoto rats (WKY) are placed on the S genetic background. Two distinct QTLs were found where one QTL was anticipated. These each accounted for a blood pressure of 15–20 mm Hg in rats fed 2% NaCl diet for 24 days. One QTL was in the <9-cM interval between D2Rat35 and D2Wox18 (Fgg), and the other was in the <7-cM interval between D2Wox18 (Fgg) and D2Mgh10. A third tentative QTL was suggested, but not clearly established, in the <3-cM interval between D2Mgh10 and D2Rat259. Received: 26 July 2001 / Accepted 6 September 2001     

High-resolution mapping of the blood pressure QTL on chromosome 7 using Dahl rat congenic strains

It was previously shown using Dahl salt-sensitive (S) and salt-resistant (R) rats that a blood pressure quantitative trait locus (QTL) was present on rat chromosome 7. In the present work, this QTL was localized to a region less than 0.54 cM in size on the linkage map using a series of congenic strains. This region was contained in a single yeast artificial chromosome that was 220 kb long. This small segment still contained the primary candidate locus Cyp11b1 (11beta-hydroxylase), but the adjacent candidate genes Cyp11b2 (aldosterone synthase) and Cyp11b3 were ruled out. It is concluded that 11beta-hydroxylase, through its known genetic variants altering the production of 18-hydroxy-11-deoxy corticosterone, is very likely to account for the blood pressure QTL on chromosome 7 in the Dahl rat model of hypertension. This QTL accounts for about 23 mm Hg under the condition of 2% NaCl diet for 24 days.     

Localization of a blood pressure QTL to a 2.4-cM interval on rat chromosome 9 using congenic strains

A blood pressure (BP) quantitative trait locus (QTL) was previously found on rat chromosome 9 using Dahl salt-sensitive (S) and Dahl salt-resistant (R) rats. A congenic strain, S.R(chr9), constructed by introgressing an R chromosomal segment into the S background, previously proved the existence of a BP QTL in a large 34.2-cM segment of chromosome 9. In the current work congenic substrains were constructed from the progenitor congenic strain, S.R(chr9). BP and heart weight comparisons between these congenic substrains and their S control localized the BP QTL to a 4.6-cM interval. Two solute carrier (Na(+)/H(+) exchanger) genes, Nhe2 and Nhe4, were excluded as candidates based on their map locations. A second iteration of congenic substrains was used to localize the QTL further to a 2.4-cM interval. Another solute carrier (Cl(-)/HCO3- exchanger) gene, Ae3, is in this reduced interval and was sequenced for both S and R strains, but no coding sequence variations were found. Ae3 mRNA was not differentially expressed in the kidney of congenic compared to S rats. Although the identity of the QTL remains unknown its map location has been reduced from an interval of 34.2 to 2.4 cM.     

Linkage map of seven polymorphic markers on rat Chromosome 18

A genetic linkage map of seven polymorphic markers was created with F₂ intercross progeny of F344/N and LEW/N rats and assigned to rat Chromosome (Chr) 18. Five of the markers described were defined by simple sequence length polymorphisms (SSLPs) associated with five genes: transthyretin (TTR), trypsin inhibitor-like protein (TILP), 2 adrenergic receptor (ADRB2), olfactory neuron-specific G protein (OLF), and gap junction protein (GJA1). One marker was defined by a restriction fragment length polymorphism (RFLP) detected with a probe for the human colony stimulating factor 1 receptor (CSF1R) gene. The D18N1R locus was defined by an anonymous DNA fragment amplified by the randomly amplified polymorphic DNA (RAPD) technique with a single short primer. These seven DNA loci formed a single genetic linkage group 30.4 cM in length with the following order: TTR-6.8 cM-D18N1R-9.1 cM-TILP-4.3 cM-CSF1R-0 cM-ADRB2-10.2 cM-OLF-0 cM-GJA1. The five SSLP markers were highly polymorphic. In a total of 13 inbred rat strains analyzed (F344/ N, LEW/N, LOU/MN, WBB1/N, WBB2/N, MR/N, MNR/N, ACI/N, SHR/N, WKY/N, BN/SsN, BUF/N, and LER/N), three to six alleles were detected for each marker. Remarkable linkage conservation was detected between the region of rat Chr 18 mapped and a region of mouse Chr 18. However, genes associated with these markers have been mapped to three different human chromosomes (Chrs 5, 6, and 18). The markers described here should be useful for genetic mapping studies and genetic monitoring of inbred rat strains.     

Defining the blood pressure QTL on Chromosome 7 in Dahl rats by a 177-kb congenic segment containing Cyp11b1

Previously we reported that there is a blood pressure quantitative trait locus (QTL) on rat Chromosome (Chr) 7 seen when comparing Dahl salt-sensitive (S) rats and Dahl salt-resistant (R) rats. Evidence was also presented that this QTL was due to genetic variants in the adrenal steroidogenic enzyme 11beta-hydroxylase ( Cyp11b1). A series of congenic strains supported this contention. In the present work we have constructed a final congenic substrain that retains a blood pressure effect and that has an introgressed congenic segment which includes Cyp11b1 and is < 177 kb in size. None of the other genes in the congenic region (eight known genes) have known biological functions for influencing blood pressure. We believe that we have reached the limit of resolution for congenic analysis of a QTL in a rodent animal model, and we conclude that Cyp11b1 causes the observed QTL on rat Chr 7 in Dahl rats.     

Congenic mapping of a blood pressure QTL on Chromosome 16 of Dahl rats

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 congenic 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.     

Improved linkage map and thirty new microsatellite markers for rat Chromosome 10

An improved linkage map for rat Chromosome (Chr) 10 with two F₂ populations was constructed. Thirty new microsatellite markers were generated from a Chr 10-specific, small-insert genomic library and mapped to rat Chr 10. Among them were the rat homologs for the mouse gene for light and heavy chains of myeloperoxidase and human neurofibromatosis 1. Eight newly generated markers (D10Mco62, D10Mco63, D10Mco64, D10Mco65, D10Mco67, D10Mco68, D10Mco70, and D10Mco74) were mapped to the region of the rat Chr 10 blood pressure QTL. The availability of such markers may be instrumental in the search for genes responsible for the hypertension. Received: 13 July 1998 / Accepted: 9 September 1998     

Human Chromosome 10 loci map to three different sheep chromosomes

The interleukin 2 receptor (IL2RA), a human Chromosome (Chr) 10p locus, was mapped to sheep Chr 13q12-q15 by in situ hybridization. Two loci from human Chr 10q, cytochrome P450 subfamily XVII (CYP17) and the tachykinin 2 receptor (TAC2R), were assigned to sheep Chrs 22q21-q23 and 25q14-q23 respectively. The assignment of IL2RA allows the provisional assignment of the previously unassigned sheep syntenic group U15 to sheep Chr 13. Sheep linkage group 5 is predicted to be located on sheep Chr 25 on the basis of the TAC2R assignment.     

Linkage map construction and QTL analysis for internal heat necrosis in autotetraploid potato

Key message

A tetraploid potato population was mapped for internal heat necrosis (IHN) using the Infinium ^® 8303 potato SNP array, and QTL for IHN were identified on chromosomes 1, 5, 9 and 12 that explained 28.21% of the variation for incidence and 25.3% of the variation for severity. This research represents a significant step forward in our understanding of IHN, and sets the stage for future research focused on testing the utility of these markers in additional breeding populations.

Abstract

Internal heat necrosis (IHN) is a significant non-pathogenic disorder of potato tubers and previous studies have identified AFLP markers linked to IHN susceptibility in the tetraploid, B2721 potato mapping population. B2721 consists of an IHN susceptible×resistant cross: Atlantic×B1829-5. We developed a next-generation SNP-based linkage map of this cross using the Infinium^® 8303 SNP array and conducted additional QTL analyses of IHN susceptibility in the B2721 population. Using SNP dosage sensitive markers, linkage maps for both parents were simultaneously analyzed. The linkage map contained 3427 SNPs and totaled 1397.68 cM. QTL were detected for IHN on chromosomes 1, 5, 9, and 12 using LOD permutation thresholds and colocation of high LOD scores across multiple years. Genetic effects were modeled for each putative QTL. Markers associated with a QTL were regressed in models of effects for IHN incidence and severity for all years. In the full model, the SNP markers were shown to have significant effects for IHN (p < 0.0001), and explained 28.21% of the variation for incidence and 25.3% of the variation for severity. We were able to utilize SNP dosage information to identify and model the effects of putative QTL, and identify SNP loci associated with IHN resistance that need to be confirmed. This research represents a significant step forward in our understanding of IHN, and sets the stage for future research focused on testing the utility of these markers in additional breeding populations.

     

Single and multiple congenic strains for hydrocephalus in the H-Tx rat

The H-Tx rat has fetal-onset hydrocephalus with a complex mode of inheritance. Previously, quantitative trait locus mapping using a backcross with Fischer F344 rats demonstrated genetic loci significantly linked to hydrocephalus on Chromosomes 10, 11, and 17. Hydrocephalus was preferentially associated with heterozygous alleles on Chrs 10 and 11 and with homozygous alleles on Chr 17. This study aimed to determine the phenotypic contribution of each locus by constructing single and multiple congenic strains. Single congenic rats were constructed using Fischer F344 as the recipient strain and a marker-assisted protocol. The homozygous strains were maintained for eight generations and the brains examined for dilated ventricles indicative for hydrocephalus. No congenic rats had severe (overt) hydrocephalus. A few pups and a significant number of adults had mild disease. The incidence was significantly higher in the C10 and C17 congenic strains than in the nonhydrocephalic F344 strain. Breeding to F344 to make F.H-Tx C10 or C11 rats heterozygous for the hydrocephalus locus failed to produce progeny with severe disease. Both bicongenic and tricongenic rats of different genotype combinations were constructed by crossing congenic rats. None had severe disease but the frequency of mild hydrocephalus in adults was similar to congenic rats and significantly higher than in the F344 strain. Rats with severe hydrocephalus were recovered in low numbers when single congenic or bicongenic rats were crossed with the parental H-Tx strain. It is concluded that the genetic and epigenetic factors contributing to severe hydrocephalus in the H-Tx strain are more complex than originally anticipated.     

Linkage of Nat and Es-1 in the mouse and development of strains congenic for N-acetyltransferase

The human polymorphism in the hepatic enzyme N-acetyltransferase (NAT) affects the rate at which individuals acetylate, and in many cases detoxify, aromatic amine and hydrazine drugs and xenobiotics. Differences in NAT activity are known to affect individual susceptibility to drug toxicities and are thought to play a part in some spontaneous disorders. A mouse model for the human acetylation polymorphism has been previously characterized and involves the A/J (slow acetylator) and C57BL/6J (rapid acetylator) inbred strains. Strain distribution analysis of 40 A x B and B x A recombinant inbred (RI) strains indicated linkage between the N-acetyltransferase gene (Nat) and the esterase 1 (Es-1) gene, located on mouse chromosome 8. A double backcross involving 107 animals confirmed the recombination frequency between Nat and Es-1 to be 12 +/- 3% (mean +/- SE). The information obtained in the backcross and RI studies was combined, yielding a 13 +/- 2.8% (mean +/- SD) recombination frequency. The Es-1 genotype was determined in our newly developed congenic strains A.B6-Natr and B6.A-Nats. The B6.A-Nats strain has the Es-1 genotype of its inbred partner, the B6 strain, and the A.B6-Natr strain has the Es-1 genotype of the donor strain. These congenic strains will be important in determining the role of the NAT genotype in susceptibility to arylamine-induced cancer and other disorders.     

Mapping the diabetes polygene Idd3 on mouse Chromosome 3 by use of novel congenic strains

Development of novel congenic mouse strains has allowed us to better define the location of the diabetogenic locus, Idd3, on Chromosome (Chr) 3. Congenic strains were identified by use of published and newly developed microsatellite markers, their genomes fingerprinted by a rapid, fluorescence-based approach, and their susceptibility to type 1 diabetes evaluated. The maximum interval containing Idd3 is now approximately 4 cM.     

Blood pressure and survival of a Chromosome 7 congenic strain bred from Dahl rats

11β-hydroxylase (Cyp11b1) mutations were previously linked to altered steroid biosynthesis and blood pressure in Dahl salt-resistant (R) and Dahl salt-sensitive (S) rats. In the present work, interval mapping identified a putative blood pressure quantitative trait locus (QTL) near Cyp11b1 in an F₁(S×R)×S population (LOD = 2.0). Congenic rats (designated S.R-Cyp11b) were constructed by introgressing the R-rat Cyp11b1 allele into the S strain. S.R-Cyp11b rats had significantly lower blood pressure and heart weight compared with S rats, proving the existence of a blood pressure QTL on Chromosome (Chr) 7 despite the fact that QTL linkage analysis of blood pressure never achieved stringent statistical criteria for significance. To test the effects of the introgressed region on blood pressure and survival, S.R.-Cyp11b and S rats were maintained on a 4% NaCl diet until they died or became moribund. Analysis of variance (ANOVA) indicated significant strain differences in blood pressure and days survived (P < 0.0001 for both) as well as gender differences in days survived (P = 0.0003). Kaplan-Meier survival analysis also found significant strain (P < 0.0001) and gender (P = 0.007) differences in days survived. However, when the effects of blood pressure were removed, significant strain differences in survival essentially disappeared. This suggests that the increased survival of S.R-Cyp11b rats was largely due to their decreased blood pressure and thus strongly corroborates the existence of a blood pressure QTL on Chr 7 near or at Cyp11b1. Received: 7 April 1997 / Accepted: 10 August 1997     

A genetic map of microsatellite markers on rat Chromosome 7

Nine microsatellite loci were mapped to rat Chromosome (Chr) 7 by genetic linkage and somatic cell hybrid analysis. These loci include the gene encoding a member of the IID sub-family of cytochrome P450 (Cyp2d), a gene with repetitive sequences expressed during myotube formation (D7Arb1e), four anonymous loci, D7Arb81, D7Arb208, D7Arb569, D7Arb609a, and three DNA loci defined by MapPair^TM markers R245, R513, and R1071. The nine loci were all identified by PCR-based microsatellite polymorphism analysis and were characterized in 40 F₂ intercross progeny of Fischer (F344/N) and Lewis (LEW/N) rats for segregation analysis. These markers formed a single linkage group spanning 76.8 cM with the following order and distances: D7Arb569-11.4 cM-D7Arb81-9.7 cM-R513-2.6 cM-Cyp2d-0.0 cM-R245-1.3 cM-D7Arb1e-10.4 cM-R1071-15.9 cM-D7Arb609a-15.4 cM-D7Arb208. Physical mapping of Cyp2d by somatic cell hybrid analysis allowed us to assign this linkage group to rat Chr 7. For each marker, two to six alleles were detected in a panel of 16 inbred rat strains (ACI/N, BN/SsN, BUF/N, DA/Bkl, F344/N, LER/N, LEW/N, LOU/MN, MNR/N, MR/N, SHR/N, SR/Jr, SS/Jr, WBB1/N, WBB2/N, WKY/N).     

Congenic mapping of a blood pressure QTL region on rat chromosome 10 using the Dahl salt-sensitive rat with introgressed alleles from the Milan normotensive strain

Multiple blood pressure (BP) quantitative trait loci (QTLs) are reported on rat chromosome 10 (RNO10). Of these, QTLs detected by contrasting the genome of the hypertensive Dahl salt-sensitive (S) rat with two different relatively normotensive strains, Lewis (LEW) and the Milan normotensive strain (MNS), are reported. Because the deduced QTL regions of both S vs. LEW and S vs. MNS comparisons are within large genomic segments encompassing more than 2 cM, there was a need to further localize these QTLs and determine whether the QTLs are unique to specific strain comparisons. Previously, the S.MNS QTL1 was mapped to less than 2.6 cM as a differential segment between two congenic strains. In this study, multiple congenic strains spanning the projected interval were studied. The BP effect of each strain was interpreted as the net effect of alleles introgressed within that congenic strain. The results suggest that the MNS alleles within the previously proposed differential segment (D10Rat27-D10Rat24) do not independently lower BP of the S rat. However, another congenic strain, S.MNS(10) × 9, containing introgressed MNS alleles that are outside of the previously proposed differential segment is of interest because (1) it demonstrated a BP-lowering effect, (2) it is contained within a single congenic strain and is not based on the observed effect of a differential segment, and, more importantly, (3) it overlaps with the previously identified S.LEW BP QTL region. Identification of the same QTL affecting BP in multiple rat strains will provide further support for the QTL’s involvement and importance in human essential hypertension.