Application of AFLP markers for QTL mapping in the rabbit.

Two rabbit (Oryctolagus cuniculus) inbred strains (AX/JU and IIIVO/JU) have been used for genetic analysis of quantitative traits related to dietary cholesterol susceptibility. Application of the AFLP (amplified fragment length polymorphism) technique with 15 primer combinations revealed 226 polymorphisms between the 2 inbred strains. A total of 57 animals from a backcross progeny (IIIVO/JU x [IIIVO/JU x AX/JU]F1) were available for the genetic analysis. These backcross animals were fed a commercial pelleted diet fortified with 0.3% w/w cholesterol during a test period that lasted five weeks. A male genetic map could be constructed, consisting of 12 linkage groups and 103 AFLP markers. Linkage analysis between the cholesterol-related traits and marker loci revealed a significant LOD score for the relative weight of adrenal glands in males (LOD score = 3.83), whereas suggestive linkages were found for basal serum total cholesterol levels in females (LOD score = 2.69), for serum total cholesterol response (area under the curve) in males (LOD score = 2.21), and for hematocrit in males (LOD score = 3.24).

Chromosomal localization of genes involved in biosynthesis, metabolism or transport of cholesterol in the rat.

Several genes involved in biosynthesis, transport or metabolism of cholesterol have been localized on rat chromosomes by using a radiation hybrid (RH) panel. The genes, coding for squalene epoxidase (Sqle), mevalonate kinase (Mvk), and farnesyl diphosphate farnesyl transferase 1 (Fdft1) which are involved in cholesterol biosynthesis, have been mapped on chromosome 7, 12, and 15, respectively. The genes coding for phospholipid transfer protein (Pltp), sterol carrier protein-2 (Scp2), ATP binding cassette reporter A7 (Abca7), scavenger receptor class B, type 1 (Cd36l1), steroidogenic acute regulatory protein (Star), and lecithin:cholesterol acyl transferase (Lcat), which are involved in the transfer and/or metabolism of cholesterol, have been mapped on chromosome 3, 5, 7, 12, 16, and 19, respectively. Each of the genes Scp2, Sqle and Fdft1 maps close to a QTL for serum total cholesterol in rat, suggesting that these three genes might represent candidate genes for the previously mapped QTLs.

A rat linkage map based on BC x LEW intercross.

A genetic linkage map consisting of 258 polymorphic loci has been constructed on the basis of an F2 intercross between the BC/CpbU and LEW/OlaHsd inbred rat strains. When compared to previously published maps a discrepancy was found for rat chromosome 7. The map spans a sex-averaged genetic length of 1790 cM and has an average marker spacing of 7.7 cM. It was estimated that this genetic map is linked to about 90% of the DNA in the rat genome. Because LEW/OlaHsd and BC/CpbU strains differ for dietary cholesterol susceptibility and hepatic copper content, the map is considered to be a valuable tool for studying the genetic background of these complex traits.

Mapping of rabbit chromosome 1 markers generated from a microsatellite-enriched chromosome-specific library.

A genomic DNA library was produced from flow-sorted rabbit chromosome 1 and enriched for fragments containing CA-repeats. Clones containing CA-repeats were identified and primers for amplification of the microsatellite were developed after sequencing the clone. The degree of polymorphism was tested in rabbits from different breeds. This approach identified 12 microsatellite markers which could be used for studying linkage relationships in the progeny of an F(2)-intercross: (AX/JUxIIIVO/JU) F(2), and two backcrosses: (OS/JxX/J)X/J and (WH/JxX/J)X/J. Seven of these markers were mapped on chromosome 1.

Sequencing and chromosomal assignment of the rat endothelial-derived lipase gene (Lipg).

Part of the nucleotide sequence of the Lipg gene in the rat was established using primers based on the mRNA sequence described in the mouse. The rat intron sequence served as a template for designing primers for the specific amplification of rat Lipg. A rat-hamster radiation hybrid (RH) panel was used for chromosomal assignment of the rat Lipg gene. The Lipg gene was found to be located on rat chromosome 18 in the vicinity of the marker D18Mit11; a region reported to be homologous with both human and mouse chromosome 18.

Genetics of a tissue esterase polymorphism (Est-6) in the rabbit (Oryctolagus cuniculus).

Genetic analysis of a polymorphic tissue esterase revealed a new locus (Est-6) with two alleles (Est-6a and Est-6b) on linkage group VI of the rabbit. Est-6 is closely linked to the Est-1,2,4 cluster. Esterase of Est-6 is found in many organs, particularly in liver and small intestine, but not in erythrocytes and serum. Est-6 esterase hydrolyzes alpha-naphthyl acetate and butyrate, naphthol AS-D acetate, indoxyl acetate, and butyrate as well as 5-bromoindoxyl acetate, N-acetyl-L-alanine-alpha-naphthyl ester but not 4-methylumbelliferyl acetate and fluorescein diacetate. The enzyme is inhibited by bis-p-nitrophenyl phosphate and eserine but not by p-chloromercuribenzoate. It was classified as a carboxylesterase (EC 3.1.1.1). Based on chromosomal localization, tissue distribution, substrate specificity, inhibitor sensitivity, and range of pI's, rabbit Est-6 is assumed to be homologous with mouse Es-7.

Genetics of two tissue esterase polymorphisms (Est-4 and Est-5) in the rabbit.

Two polymorphic esterase systems were found after electrophoresis of rabbit tissue homogenates. Each of these systems is controlled by an autosomal locus with two alleles. Est-4 determines the absence (Est-4a) or presence (Est-4b) of two bands of esterase activity with intermediate anodcal mobility and broad substrate specificity. This polymorphism was found to be present in liver, small intestine, and spleen but not in kidney, heart, and testis. Est-5 is coding for cathodally migrating esterases which differ in mobility (Est-5a and Est-5b). This polymorphism was found only in kidney and testis homogenates. Est-5 esterases are more active against alpha-naphthyl acetate than against beta-naphthyl acetate and have no activity against alpha-naphthyl butyrate. Linkage analysis indicated that Est-4 is localized on rabbit LG VI as part of a cluster of esterase loci, whereas Est-5 segregates independently, Rabbits from two inbred and nine partly inbred strains were tested for these polymorphisms.

Genetic control of plasma cholesterol response in the rat.

The inheritance of the susceptibility for dietary cholesterol in the rat has been studied by testing the plasma cholesterol response in male animals from crosses between hyperresponding and hyporesponding inbred strains. Comparison of the variances of the response of genetically uniform groups (parental strains and F1 hybrid) and segregating groups (backcrosses and F2 hybrid) revealed that more than 80 percent of the observed variation can be attributed to additive genetic factors and indicated that two major genes are involved in the control of the plasma cholesterol response.

Separation of beta-D-galactosidases in rabbit tissues: genetics of neutral beta-D-galactosidase.

Three different types of beta-D-galactosidase (EC 3.2.1.23) could be distinguished in rabbit tissues using electrophoretic procedures. (1) Acid beta-D-galactosidase with a low mobility and maximal activity at pH 3-5 was found in the particulate fraction of various tissue homogenates. This enzyme hydrolyzed 4-methylumbelliferyl-D-galactoside, but no activity against other glycoside substrates could be demonstrated. The enzyme was inhibited by galactono-(1 leads to 4)-lactone. (2) Lactose-hydrolyzing beta-D-galactosidase with an intermediate mobility was found only in juvenile small intestine. Most of the activity was found in the particulate fraction of the cell. The enzyme hydrolyzed several other synthetic glycoside substrates besides lactose. It was most active at pH 5-6 and strongly inhibited by glucono-(1 leads to 5)-lactone but not much affected by galactono-(1 leads to 4)-lactone. (3) Neutral beta-D-galactosidase with a fast mobility and maximal activity at pH 6-8 was found in the soluble fraction of homogenates from liver, kidney, and small intestine. This enzyme also showed a broad substrate specificity; it possessed activity against aryl-beta-D-glucoside, -fucoside, and -galactoside substrates but not against lactose. The enzyme was strongly inhibited by glucono-(1 leads to 5)-lactone and (less) by galactone-(1 leads to 4)-lactone. Neutral beta-D-galactosidase and neutral beta-D-glucosidase (EC 3.2.1.21) are probably identical enzymes in the rabbit. Individual variation, in both electrophoretic mobility and activity, was found for neutral beta-D-galactosidase. Genetic analysis of the electrophoretic variants revealed that two alleles at an autosomal locus are responsible for this variation.

Cholesterol response in inbred strains of rats, Rattus norvegicus.

A hypercholesterolemic diet fed to rats revealed significant interstrain differences in plasma cholesterol levels. Hyperresponding and hyporesponding strains could be distinguished from normoresponding strains within 3 weeks. The increase in plasma cholesterol level was more than 300 mg/100 ml in the hyperresponding strains BN/Cpb, SD/Cpb and WE/Z and less than 50 mg/100 ml in the hyporesponding strains S3/Cpb and SHR/Cpb. These differences were primarily genetically determined: the calculated coefficient of genetic determination (g2) of the response was 0.84. The response is not correlated with variation in plasma esterase or alkaline phosphatase isozyme patterns.

Biochemical polymorphism in the rat: genetics of three electrophoretic variants and characterization of inbred strains.

Nine inbred strains of the rat (Rattus norvegicus) were screened for differences in electrophoretically detectable proteins. Interstrain variation was observed for 7 of 26 proteins. Three of these variants have not been described previously: leucine aminopeptidase (Lap-1), major urinary protein (Mup-1), and seminal vesicle protein (Svp-2). Genetic analysis revealed two autosomal alleles for each of these polymorphisms. The loci Lap=1, Mup-1, and Svp-2 are linked neither to one another nor to the previously described Svp-1 and Es-4 loci. Each of the nine strains can be identified now by a specific set of monogenic markers.

Genetic and physiological aspects of cholesterol accumulation in hyperresponding and hyporesponding rabbits.

After a 5 week period of feeding a cholesterol-rich diet to rabbits, hyperresponders with high plasma cholesterol levels and hyporesponders with low plasma cholesterol levels could be distinguished from normal responders. The response was found to be correlated with the esterase genotype at the Est-2 locus. The increase in total body cholesterol was higher in hyper-than in hyporesponders. In both groups most of the accumulated dietary cholesterol was found in plasma and liver. Adrenal weight and plasma corticosterone levels were more increased in hyper- than in hyporesponders. The cholesterol-rich diet resulted in an augmentation of liver lipase and lipoprotein lipase activities. These lipolytic activities were more increased in hyper- than in the hyporesponders.

Serum esterase genetics in rabbits. IV. The prealbumin and beta-globulin systems.

Discontinuous starch gel electrophoresis revealed a fourth allele of rabbit pre-albumin serum esterase at locus Est-2. This allele is designated Est-2f and appears to be silent. In addition to the prealbumin serum esterases, another serum esterase system was studied in rabbits. This system is localized in the beta-globulin region. Genetic analysis indicated that one locus with two codominant alleles controls the variation in this region. Linkage of this system with Est-1 and Est-2 of the prealbumin serum esterases was demonstrated. Comparison of the arrangement of these esterase loci on linkage group VI with the esterase loci on chromosome 8 of the mouse gives additional support for the theory of evolutionary conservation of chromosomal segments coding for mammalian esterases.

Serum esterase genetics in rabbits. III. A third allele on the Est-2 locus.

A fast-migrating F' zone of the prealbumin serum esterase system of rabbits is demonstrated in low frequency in the breed Vienna White (stock Cpb:VW). Evidence is given that this zone is controlled by a third allele of the Est-2 locus. The zymotypic expression of this allele (Est-2-F') shows codominance in combination with the Est-2-F allele and complete dominance in combination with the Est-2-F' allele. In contradistinction to the F zones of the Est-2-F allele, the F' zone possesses no atropinesterase activity.