RESUMO
Livestock guarding dogs are a valuable adjunct to the pastoral community. Having been traditionally selected for their working ability, they fulfil their function with minimal interaction or command from their human owners. In this study, the population structure and the genetic differentiation of three Italian livestock guardian breeds (Sila's Dog, Maremma and Abruzzese Sheepdog and Mannara's Dog) and three functionally and physically similar breeds (Cane Corso, Central Asian Shepherd Dog and Caucasian Shepherd Dog), totalling 179 dogs unrelated at the second generation, were investigated with 18 autosomal microsatellite markers. Values for the number of alleles per locus, observed and expected heterozygosity, Hardy-Weinberg Equilibrium, F stats, Nei's and Reynold's genetic distances, clustering and sub-population formation abilities and individual genetic structures were calculated. Our results show clear breed differentiation, whereby all the considered breeds show reasonable genetic variability despite small population sizes and variable selection schemes. These results provide meaningful data to stakeholders in specific breed and environmental conservation programmes.
Assuntos
Cruzamento , Cães , Variação Genética , Animais , Cães/genética , Itália , Gado , Repetições de MicrossatélitesRESUMO
The possibility of using genetic control strategies to increase disease resistance to infectious diseases relies on the identification of markers to include in the breeding plans. Possible incomplete exposure of mastitis-free (control) animals, however, is a major issue to find relevant markers in genetic association studies for infectious diseases. Usually, designs based on elite dairy sires are used in association studies, but an epidemiological case-control strategy, based on cows repeatedly field-tested could be an alternative for disease traits. To test this hypothesis, genetic association results obtained in the present work from a cohort of Italian Holstein cows tested for mastitis over time were compared with those from a previous genome-wide scan on Italian Holstein sires genotyped with 50k single nucleotide polymorphisms for de-regressed estimated breeding values for somatic cell counts (SCCs) on Bos taurus autosome (BTA6) and BTA14. A total of 1121 cows were selected for the case-control approach (cases=550, controls=571), on a combination of herd level of SCC incidence and of within herd individual level of SCC. The association study was conducted on nine previously identified markers, six on BTA6 and four on BTA14, using the R statistical environment with the 'qtscore' function of the GenABEL package, on high/low adjusted linear score as a binomial trait. The results obtained in the cow cohort selected on epidemiological information were in agreement with those obtained from the previous sire genome-wide association study (GWAS). Six out of the nine markers showed significant association, four on BTA14 (rs109146371, rs109234250, rs109421300, rs109162116) and two on BTA6 (rs110527224 and rs42766480). Most importantly, using mastitis as a case study, the current work further validated the alternative use of historical field disease data in case-control designs for genetic analysis of infectious diseases in livestock.
Assuntos
Estudos de Associação Genética/veterinária , Mastite Bovina/genética , Animais , Estudos de Casos e Controles , Bovinos , Feminino , Estudos de Associação Genética/métodos , Itália , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Accurate pedigrees are essential to optimize genetic improvement and conservation of animal genetic resources. In goats, the use of mating groups and kidding management procedures hamper the identification of parentage. Small panels of single nucleotide polymorphisms (SNP) have been proposed in other species to substitute microsatellites for parentage assessment. Using data from the current GoatSNP50 chip, we developed a new 3-step procedure to identify a low-density SNP panel for highly accurate parentage assessment. Methodologies for SNP selection used in other species are less suitable in the goat because of uncertainties in the genome assembly. The procedure developed in this study is based on parent-offspring identification and on estimation of Mendelian errors, followed by canonical discriminant analysis identification and stepwise regression reduction. Starting from a reference sample of 109 Alpine goats with known pedigree relationships, we first identified a panel of 200 SNP that was further reduced to 2 final panels of 130 and 114 SNP with random coincidental match inclusion of 1.51×10(-57) and 2.94×10(-34), respectively. In our reference data set, all panels correctly identified all parent-offspring combinations, revealing a 40% pedigree error rate in the information provided by breeders. All reference trios were confirmed by official tests based on microsatellites. Panels were also tested on Saanen and Teramana breeds. Although the testing on a larger set of breeds in the reference population is still needed to validate these results, our findings suggest that our procedure could identify SNP panels for accurate parentage assessment in goats or in other species with unreliable marker positioning.
Assuntos
Cabras/genética , Polimorfismo de Nucleotídeo Único , Animais , Cruzamento , Repetições de Microssatélites , LinhagemRESUMO
Genetic variation at the αS1-casein locus (CSN1S1) is recognized as being crucial in the selection of dairy goats for cheese yield. At this locus, the existence of alleles that have strong, intermediate, weak, and null favorable effects on cheese yield and curd firmness is well known. Selection for alleles that have a strong favorable effect has been deliberately carried out, especially in France. In fact, the importance of αS1-casein in selection was recently confirmed in the selling policies of semen, where bucks are marketed according to their genotypes. We evaluated genotypes and alleles frequencies at the αS1-casein locus in 491 Italian Saanen and Alpine goats and compared them with previous data to investigate their evolution over the past decade. We also estimated soft cheese yield in a subset of the most represented genotypes to quantify the economic importance of considering the genetic trend of αS1-casein genotype frequencies. We found a significant increase in frequency of the allele with the strongest favorable effect, A (+12 and +13%), and of the intermediate allele E (+17 and +7%) in Saanen and Alpine goats, respectively. Surprisingly, the frequency of the strong allele B decreased strikingly over time (-12% in Saanen, -6% in Alpine from 2004 to 2012). This is consistent with the current marketing of semen, in that bucks that are homozygous for strong (AA and BB) and intermediate alleles (EE) and even heterozygous for these alleles (BE and AE) are considered equal. It is worth noting that this practice strongly penalizes the best breeders that have flocks composed almost entirely of goats that are homozygous for strong alleles. For heterozygous goats, we estimated an economic loss of 85 and 215 per goat per lactation, respectively, for AE and BE, compare with AA and BB genotypes. The marketing of buck semen should clearly differentiate these 2 alleles to ensure the best economic genetic progress at this locus.
Assuntos
Caseínas/genética , Queijo/análise , Cabras/genética , Leite/metabolismo , Alelos , Animais , Cruzamento , Feminino , Frequência do Gene , Loci Gênicos , Genótipo , Cabras/fisiologia , Lactação , MasculinoRESUMO
Milk yield and composition are of great economic importance for the dairy goat industry. The identification of genes associated with phenotypic differences for these traits could allow for the implementation of gene-assisted selection programs in goats. Associations between polymorphisms at 3 candidate genes and milk production traits in Alpine goats farmed in Italy were investigated in the present research. Considered genes were acetyl-coenzyme A carboxylase α (ACACA), the major regulatory enzyme of fatty acid biosynthesis; stearoyl-coenzyme A desaturase (SCD), involved in the biosynthesis of monounsaturated fatty acids in the mammary gland; and lipoprotein lipase (LPL), which plays a central role in plasma triglyceride metabolism. An approach somewhat similar to the granddaughter design for detecting quantitative trait loci in dairy cattle was followed. Effects of genotypes of a sample of 59 Alpine bucks on phenotypes of their 946 daughters raised in 75 flocks were investigated. Data comprised 13,331 daily records for milk yields (L/d), fat and protein yields (kg/d), and fat and protein contents (%) of 2,200 lactations. Population genetics parameters were calculated and associations between milk production traits and 10 single nucleotide polymorphisms (SNP) at the 3 genes were tested. Two markers at the ACACA, 1 for the SCD and 1 at the LPL locus, deviated significantly from the Hardy-Weinberg equilibrium, with an observed heterozygosity lower than expected. Flock, age of the goat, kidding season, and stage of lactation affected all traits considered, except fat percentage. Three SNP were found to be significantly associated with milk production traits. The SNP located on the ACACA gene showed an effect on milk yield, with daughters of TT bucks having an average test-day milk yield of about 0.3 to 0.25 L/d lower than the other 2 genotypes. The marker on the LPL locus was highly associated with milk yield, with the largest values for CC daughters (about 0.50L more than GG). The TGT deletion located on the untranslated region of the SCD gene showed significant effects on average milk and protein yields. The homozygote-deleted genotype had values about 0.5 L/d and 16 g/d lower for milk and protein daily yield, respectively, compared with the TGT/TGT genotype. Differences between genotypes were quite constant across most of the lactation. Associations found in the present study, which should be tested in a larger sample, especially for those markers that show rare genotypes, may offer useful indications for the genetic improvement of dairy traits in goats.
Assuntos
Acetil-CoA Carboxilase/genética , Cabras/genética , Lactação/genética , Lipase Lipoproteica/genética , Estearoil-CoA Dessaturase/genética , Acetil-CoA Carboxilase/fisiologia , Alelos , Animais , Gorduras/análise , Feminino , Estudos de Associação Genética/veterinária , Genótipo , Cabras/metabolismo , Cabras/fisiologia , Lactação/fisiologia , Lipase Lipoproteica/fisiologia , Masculino , Leite/química , Proteínas do Leite/análise , Polimorfismo de Nucleotídeo Único/genética , Estearoil-CoA Dessaturase/fisiologiaRESUMO
Associations of allelic variants of the ß-lactoglobulin (LGB), stearoyl-coenzyme A desaturase 1 (SCD), and sterol regulatory element binding protein (SREBP-1) genes with milk production, composition (fat, protein, and casein content), acidity (pH and titratable acidity), and coagulation properties (rennet coagulation time and curd firmness) were investigated in Brown Swiss cows. In total, 294 animals (progeny of 15 sires) reared in 16 herds were milk-sampled once. The additive effects of LGB (rs109625649:C>T), SCD (ss469414220:C>T), and SREBP-1 (AB355704: g.101_185ins) polymorphisms on the aforementioned traits were analyzed through Bayesian linear models. The LGB genotype affected rennet coagulation time, with TT (or BB) alleles showing longer rennet coagulation time compared with CC (or AA) cows. The SCD gene allelic variants were found to be associated with protein and casein contents and curd firmness: CC animals had the lowest values for the aforementioned traits. An association was found between SREBP-1 alleles and fat content, with the highest values for cows carrying the 84-bp insertion (or L) allele. Results suggest a possible use of these loci in gene-assisted selection programs for the improvement of milk quality traits and coagulation properties in Brown Swiss cattle.
Assuntos
Lactação/genética , Lactoglobulinas/genética , Leite/química , Estearoil-CoA Dessaturase/genética , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Alelos , Animais , Bovinos/genética , Feminino , Estudos de Associação Genética/veterinária , Variação Genética , Genótipo , Concentração de Íons de Hidrogênio , Leite/metabolismo , Proteínas do Leite/análise , Polimorfismo Genético/genéticaRESUMO
Cow milk allergy is the most frequent allergy in the first years of life. Milk from other mammalian species has been suggested as a possible nutritional alternative to cow milk, but in several cases, the clinical studies showed a high risk of cross-reactivity with cow milk. In the goat species, αS1-casein (αS1-CN), coded by the CSN1S1 gene, is characterized by extensive qualitative and quantitative polymorphisms. Some alleles are associated with null (i.e., CSN1S1 0(1)) or reduced (i.e., CSN1S1 F) expression of the specific protein. The aim of this work was to obtain new information on goat milk and to evaluate its suitability for allergic subjects, depending on the genetic variation at αs1-CN. Individual milk samples from 25 goats with different CSN1S1 genotypes were analyzed by sodium dodecyl sulfate PAGE and immunoblotting, using monoclonal antibodies specific for bovine α-CN and sera from children allergic to cow milk. A lower reaction was observed to 2 goat milk samples characterized by the CSN1S1 0(1)0(1) and 0(1)F genotypes. Moreover, a fresh food skin prick test, carried out on 6 allergic children, showed the lack of positive reaction to the 0(1)0(1) milk sample and only one weak reactivity to the 0(1)F sample. The risk of cross-reactivity between cow and goat milk proteins suggests the need for caution before using goat milk for infant formulas. However, we hypothesize that it can be used successfully in the preparation of modified formulas for selected groups of allergic patients. The importance of taking the individual goat CN genetic variation into account in further experimental studies is evident from the results of the present work.
Assuntos
Caseínas/genética , Cabras/genética , Hipersensibilidade a Leite/genética , Animais , Caseínas/efeitos adversos , Bovinos , Criança , Genótipo , Humanos , Leite/química , Polimorfismo GenéticoRESUMO
Suppressive subtractive hybridization (SSH) was used to analyse the muscle transcriptome and identify genes affecting meat quality within an Italian pig population of Large White and Landrace purebred individuals. Seven phenotypes were recorded at slaughter: dorsal fat thickness, ham fat thickness, ham fat coverage, muscle compactness, marbling, meat colour and colour uniformity. Two subtractive libraries were created from longissimus dorsi tissue of selected pigs with extreme phenotypes for meat quality. Eighty-four differentially expressed ESTs were identified, which showed homology to expressed pig sequences and/or to genomic pig sequences produced within the pig genome project. Sixty-eight sequences were mapped on the pig genome, and most of these sequences co-localized with the same chromosomal positions as QTLs that have been previously identified for meat quality. Thirty sequences, including eight matching known genes previously related to muscle metabolic pathways, were selected to statistically validate their differential expression. Association analysis and t-test results indicated that 28 ESTs of the 30 analysed were associated with phenotypes investigated here and have significant differential expression levels (P≤ 0.05) between the two tails of the phenotypic distribution.
Assuntos
Regulação da Expressão Gênica/genética , Genoma/genética , Carne/análise , Técnicas de Hibridização Subtrativa/métodos , Suínos/genética , Transcriptoma , Animais , Sequência de Bases , Mapeamento Cromossômico/veterinária , Feminino , Perfilação da Expressão Gênica/veterinária , Biblioteca Gênica , Masculino , Dados de Sequência Molecular , Músculo Esquelético/metabolismo , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Análise de Sequência de DNA/veterinária , Especificidade da Espécie , Técnicas de Hibridização Subtrativa/veterinária , Suínos/metabolismoRESUMO
Several lipogenic genes have been shown to have effects on lipid metabolism: stearoyl CoA desaturase 1 (SCD1) catalyzes the desaturation of several fatty acids (FA) in the cis-Delta(9) position in mammary glands of ruminant animals, diacylglycerol acyltransferase 1 (DGAT1) is a key enzyme in triacylglycerol synthesis in the mammary gland, and sterol regulatory element binding protein (SREBP-1) is a transcription factor that regulates expression levels of the SCD1 gene and other genes relevant to lipid and FA metabolism in adipose tissue and mammary gland. In this work, 351 Italian Brown cows were genotyped for polymorphisms in the SCD1, SREBP-1, and DGAT1 genes to reveal the allelic distribution in the population. Subsequently, effects on individual milk FA composition and on cis-9 unsaturated/saturated FA ratios, a proxy of mammary stearoyl CoA desaturase activity, were investigated. The genotypes of SCD1 (A293V) and DGAT1 (K232A) were determined by an approach based on the ligation detection reaction and a universal array, whereas the genotype of SREBP-1 (84-bp insertion-deletion) was revealed by PCR amplification of intron 5. The genotype analysis showed an unbalanced distribution of alleles within all genes, being the allele with higher gene frequency at 82, 84, and 98% for SCD1, SREBP-1, and DGAT1, respectively. Significant associations between SCD1 and DGAT1 polymorphisms and milk FA composition were found, whereas SREBP-1 polymorphism was not associated with milk FA composition. In particular, SCD1 showed significant association with C14:1 cis-9 and C14:1 cis-9/C14:0, which is considered the best proxy of the desaturation activity in mammary gland. The DGAT1 polymorphism had the strongest association with milk FA composition, which confirmed the key role of DGAT1 in lipid metabolism of mammary gland. However, the unbalanced distribution of alleles in all polymorphisms investigated suggested that the size of population should be increased to confirm the results of the present study.
Assuntos
Bovinos/genética , Diacilglicerol O-Aciltransferase/genética , Ácidos Graxos/análise , Leite/química , Polimorfismo Genético/genética , Estearoil-CoA Dessaturase/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Animais , Sequência de Bases , Feminino , Dados de Sequência Molecular , Alinhamento de SequênciaRESUMO
The suppressive subtractive hybridization technique was previously used by the authors to identify candidate genes for meat quality in pig. A set of ESTs homologous (>95%) to genes involved in muscle metabolism is reported in the present paper. Four ESTs homologous to MYH1, KALRN, MLC2V, and SNX13 genes plus two genes (AK1, PPIA) used as housekeeping for muscle tissue were assigned to porcine chromosomes using the INRA-Minnesota 7000 rads radiation hybrid panel (IMpRH). Our data confirm and refine the cytogenetic position of the KALRN, AK1, PPIA genes, improve the existing physical map of MYH1 and assign two new genes (MLC2V and SNX13) to swine chromosomes.
Assuntos
Adenilato Quinase/genética , Ciclofilina A/genética , Proteínas Ativadoras de GTPase/genética , Isoenzimas/genética , Cadeias Pesadas de Miosina/genética , Cadeias Leves de Miosina/genética , Proteínas Serina-Treonina Quinases/genética , Suínos/genética , Animais , Mapeamento Cromossômico , Cromossomos/genética , Etiquetas de Sequências Expressas , Ligação Genética , Genoma , Mapeamento de Híbridos Radioativos/métodosRESUMO
The aim of the study was to estimate the effect of the composite CSN2 and CSN3 genotypes on milk coagulation, quality, and yield traits in Italian Holstein cows. A total of 1,042 multiparous Holstein cows reared on 34 commercial dairy herds were sampled once, concurrently with monthly herd milk recording. The data included the following traits: milk coagulation time; curd firmness; pH and titratable acidity; fat, protein, and casein contents; somatic cell score; and daily milk, fat, and protein yields. A single-trait animal model was assumed with fixed effects of herd, days in milk, parity, composite casein genotype of CSN2 and CSN3 (CSN2-CSN3), and random additive genetic effect of an animal. The composite genotype of CSN2-CSN3 showed a strong effect on both milk coagulation traits and milk and protein yields, but not on fat and protein contents and other milk quality traits. For coagulation time, the best CSN2-CSN3 genotypes were those with at least one B allele in both the CSN2 and CSN3 loci. The CSN3 locus was associated more strongly with milk coagulation traits, whereas the CSN2 locus was associated more with milk and protein yields. However, because of the tight linkage between the 2 loci, the composite genotypes, or haplotypes, are more appropriate than the single-locus genotypes if they were considered for use in selection.
Assuntos
Caseínas/genética , Bovinos/genética , Lactação/genética , Leite/química , Leite/normas , Animais , Feminino , Genótipo , Itália , Leite/metabolismo , Proteínas do Leite/análiseRESUMO
Several single nucleotide polymorphisms have been identified in the goat milk casein genes, most of them modifying the amino acid sequence of the coded protein. At least 9 variants have been found in goat beta-CN (CSN2); 6 of them were characterized at the DNA level (A, A1, C, E, 0, and 0'), whereas the other 3 variants were described only at the protein level. The recently identified silent A1 allele is characterized by a C-->T transition at the 180th nucleotide of the ninth exon. In the present work, typing results from different breeds (3 Italian, 3 German, and a composite of African breeds for a total of 335 samples) demonstrated that the same mutation is carried by the CSN2*C allele. In addition, the T nucleotide at the 180th nucleotide of the ninth exon was always associated with CSN2*C in all the breeds analyzed. Thus, another silent allele occurs at goat CSN2 and can be named CSN2*C1. The much wider distribution of C1 with respect to the A1 allele indicates that the single nucleotide polymorphisms characterizing the silent mutation originated from CSN2*C. A method for the identification of this allele simultaneously with 5 of the 6 DNA-characterized alleles is also proposed. The mutation involved codifies for the same protein of the C allele; nevertheless, its location in the 3' untranslated region of the gene might affect the specific casein expression.
Assuntos
Alelos , Cruzamento , Caseínas/genética , Cabras/genética , Animais , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Associations between stearoyl-CoA desaturase (SCD) gene polymorphisms and milk production traits (milk, fat, and protein yields, fat and protein contents, somatic cell score) were investigated on a sample of 701 lactations of 313 Italian Holsteins. Test-day records (5,097) were analyzed with a mixed linear model that included the fixed effects of herd, date of test, parity, genotype at the SCD locus, and lactation interval nested within SCD genotype, and the random effect of cow. An effect of the SCD genotype on milk and protein yields was detected, with VV cows producing more milk (about 2 kg/d) and protein (about 0.07 kg/d) compared with AA cows. The contribution of the SCD locus to the phenotypic variance of the 2 traits was about 0.015. These results suggest a possible use of the SCD locus in gene-assisted selection programs for the improvement of milk production traits in dairy cattle, although large-scale studies in different breeds are required.
Assuntos
Cruzamento , Bovinos/genética , Lactação/genética , Leite/metabolismo , Polimorfismo de Nucleotídeo Único , Estearoil-CoA Dessaturase/genética , Substituição de Aminoácidos , Animais , Gorduras/análise , Feminino , Genótipo , Hereditariedade , Itália , Análise dos Mínimos Quadrados , Masculino , Leite/química , Leite/citologia , Proteínas do Leite/análise , FenótipoRESUMO
Milk fatty acid composition is a parameter of great interest for evaluation of nutritional quality of milk. Stearoyl-CoA desaturase (SCD) is a key enzyme in mammary lipid metabolism because it is able to add a double bond in the cis delta9-position in a large spectrum of medium- and long-chain fatty acids. A polymorphism with 2 alleles (A and V) in the fifth exon of the SCD gene has been reported. The effect of SCD genotype on individual milk fatty acid composition and on cis-9 unsaturated/saturated fatty acid ratios of 297 Holstein Italian Friesian cows was investigated in this paper. The SCD genotypes were determined by using a single strand conformation polymorphism method. Relative frequencies of SCD genotypes were 27, 60, and 13% for AA, AV, and VV, respectively. Milk of AA cows had a greater content of cis-9 C18:1 and total monounsaturated fatty acids and a higher C14:1/C14 ratio than did milk of VV cows. The relative contribution of SCD genotype to variation of monounsaturated fatty acids, cis-9 C18:1, and cis-9 C14:1 was 5, 4, and 7.7%, respectively. No significant differences were detected between SCD genotypes in the milk content of cis-9, trans-11 C18:2. Results of the present work provide some indication of an association between SCD locus and the fatty acid profile in the examined sample of Italian Holsteins, thus suggesting a possible role of this gene in the genetic variation of milk nutritional properties.
Assuntos
Bovinos/genética , Ácidos Graxos/análise , Leite/química , Polimorfismo Genético , Estearoil-CoA Dessaturase/genética , Animais , Bovinos/metabolismo , DNA/análise , Gorduras/análise , Feminino , Genótipo , Itália , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Análise de Sequência de DNARESUMO
The aim of this work was to study the effects of isoelectrofocusing (IEF) milk protein variants on milk composition in the Italian Orobica goat breed, which is characterized by a rather high frequency of the kappa-casein (CSN3) B(IEF) allele. Significant associations were found between the IEF phenotype and protein and casein percentages. A favorable effect of the CSN3 B(IEF) variant was found for both protein and casein percentages, with a codominance trend for the 3 phenotypes: BB > AB > AA. Depending on the selection purpose, emphasis could be given to different kappa-casein variants in breeding. The high frequency of B(IEF) could be exploited in breeding strategies to improve the protein and casein percentages when cheese making is a selection objective.
Assuntos
Caseínas/química , Cabras/genética , Leite/química , Polimorfismo Genético , Alelos , Animais , Feminino , Focalização Isoelétrica/veterinária , Itália , Análise dos Mínimos Quadrados , Modelos Lineares , Fenótipo , Análise de Sequência de ProteínaRESUMO
The objective of this study was to develop and validate a fast method for typing the main mutations of bovine milk protein genes by using microarray technology. An approach based on the ligation detection reaction (LDR) and a universal array (UA) was used. Polymorphisms in both the coding and noncoding sequences of alpha(S1)-casein, beta-casein, kappa-casein, and beta-lactoglobulin genes were considered because of their well-known effects on milk composition and cheese production. A total of 22 polymorphic sites, corresponding to 21 different variants, were included in the diagnostic microarray. First, a multiplex PCR was developed to amplify all the DNA target sequences simultaneously. Second, the LDR-UA assay was implemented. The method was validated by analyzing 100 Italian Friesian DNA samples, which were also genotyped by conventional methods both at the protein level by means of milk isoelectrofocusing and at the molecular level using PCR-RFLP and PCR-single strand conformation polymorphism techniques. The genotypes obtained using the LDR-UA approach were in full agreement with those obtained by the conventional analyses. An important result of the LDR-UA assay was a more accurate genotyping of the different milk protein alleles than was found with conventional typing methods. At the kappa-casein gene, in fact, 4 samples were heterozygous (3 reference samples and 1 validation sample) for an allele coding for Thr(136) and Ala(148). This variant, which can be considered as the wild type of the genus Bos, is not usually identifiable by the conventional typing methods used. The multiplex PCR-LDR-UA approach developed provides for an accurate, inexpensive, and high-throughput assay that does not exhibit false positive or false negative signals, thus making it highly suitable for animal genotyping.
Assuntos
Bovinos/genética , Análise em Microsséries/veterinária , Proteínas do Leite/genética , Polimorfismo de Nucleotídeo Único/genética , Animais , Genótipo , Análise em Microsséries/métodos , Reação em Cadeia da Polimerase , Reprodutibilidade dos TestesRESUMO
The analysis of casein polymorphisms in goat species is rather difficult, because of a large number of mutations at each locus, and the tight linkage involving the 4 casein genes. Three goat breeds from Northern Italy, Orobica, Verzasca, and Frisa, were analyzed at the casein complex by milk isoelectrofocusing and analyses at the DNA level to identify the majority of all known polymorphisms. The casein gene structure of the 3 local breeds at alpha(S1)-casein (CSN1S1), beta-casein (CSN2), alpha(S2)-casein (CSN1S2), and kappa-casein (CSN3) was compared with that of Camosciata, a more widely distributed breed. A new allele was identified and characterized at CSN2 gene, which seemed to be specific to the Frisa breed. It was named CSN2*E, and was characterized by a transversion TCT --> TAT responsible for the amino acid exchange Ser(166) --> Tyr(166) in the mature protein. The casein haplotype structure is highly different among breeds. A total of 26 haplotypes showed a frequency higher than 0.01 in at least 1 of the 4 breeds considered, with 12, 3, 5, and 19 haplotypes in Frisa, Orobica, Verzasca, and Camosciata breeds, respectively. Only 13 haplotypes occurred at a frequency higher than 0.05 in at least 1 breed. With the molecular knowledge of each locus, the ancestral haplotype coding for CSN1S1*B, CSN2*A, CSN1S2*A, and CSN3*B protein variants can be postulated. A protein evolutionary model considering the whole casein haplotype is proposed.
Assuntos
Caseínas/genética , Cabras/genética , Polimorfismo Genético/genética , Alelos , Animais , Cruzamento , DNA/análise , Feminino , Frequência do Gene , Haplótipos , Focalização Isoelétrica , Itália , Desequilíbrio de Ligação , Leite/química , Filogenia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo Conformacional de Fita SimplesRESUMO
The objective of this study was to estimate the effects of different haplotypes of the casein genes on milk production traits in Italian dairy cattle. Traits of interest were yields of milk, fat, and protein, and percentages of fat and protein in milk. The data included 728 multiparous records from 347 Holsteins and 773 records from 298 Brown Swiss cows. Records were preadjusted for effects of age and parity, season of calving, and region, and expressed as deviations from herdmate averages. Twenty half-sib families were represented in each breed. Haplotype probabilities were estimated for each animal and phenotypes were regressed on these probabilities. Nine haplotypes were observed in Holsteins and 17 were identified among the Brown Swiss. For Holsteins, significant effects were observed for protein percentage, with some indication of an effect for fat percentage. For the Brown Swiss, effects of haplotypes were significant for milk yield and fat and protein percentages. Effects were strongest for protein percentage. Correlation coefficients of solutions across breeds tended to be strong and positive, indicating that the same haplotypes had similar estimated effects in the 2 breeds. Although the data were limited (<350 cows in each study), this latter result may suggest that genes in the casein complex itself are responsible for the effects observed, rather than loci that are physically linked on either side of the casein cluster.
Assuntos
Caseínas/genética , Bovinos/genética , Haplótipos/genética , Lactação/genética , Leite/química , Animais , Cruzamento , Gorduras/análise , Feminino , Lactação/fisiologia , Masculino , Leite/metabolismo , Proteínas do Leite/análise , Proteínas do Leite/genéticaRESUMO
The objective of this work was to propose an algorithm (HAPROB) to estimate haplotype probabilities for genotyped members of half-sib families for which parents lacked genotypic information. The algorithm had 2 basic steps. First, a Monte Carlo-based approach was used to estimate haplotype probabilities for sires conditional upon offspring genotypes and population allelic frequencies, and then offspring-haplotype probabilities were estimated conditional upon sire probabilities and population frequencies. The 2 steps were alternated iteratively until estimates of population frequencies were essentially unchanged. Simulation was used to evaluate effects of the number of Monte Carlo cycles on the accuracy of the reconstructed haplotypes. Fifty thousand cycles was found to be sufficient for the haplotype configurations considered. Accuracy of the algorithm was compared with that obtained by the public domain SIMWALK2 software. Predictions of the most likely haplotype configurations are produced by SIM-WALK2, but no estimates of probability are given. The accuracy of the current approach was comparable to that obtained from SIMWALK2. The proportions of times that haplotypes were reconstructed correctly were 87.0 and 92.4% (sires and offspring) for HAPROB vs. 87.5 and 91.5% for SIMWALK2. Effects of family size on accuracy of reconstruction were examined. Accuracy of reconstruction was only about 4% for sires with 2 offspring, but accuracy among the offspring themselves was 65%. Accuracy increased quickly as family size increased and reached 100% for sires with 30 offspring. Maximum accuracy for offspring was about 96%. Estimates of haplotype probabilities produced can be used in regression analyses to estimate effects of haplotypes on quantitative phenotypes.
Assuntos
Bovinos/genética , Ligação Genética/genética , Haplótipos/genética , Método de Monte Carlo , Algoritmos , Animais , Simulação por Computador , Feminino , Frequência do Gene , Funções Verossimilhança , Masculino , Modelos Genéticos , Sensibilidade e Especificidade , SoftwareRESUMO
The objectives of this study were to develop and simulate the implementation of several strategies for repeated application of quantitative trait loci (QTL) detection and marker-assisted selection (MAS) and to compare the short-term and continual genetic responses. A finite locus model was simulated with 20 QTL randomly distributed across 30 chromosome. Three hundred markers were evenly spaced across the genome. Allelic effects were sampled from a double exponential distribution. A daughter design was used every generation to determine the marker alleles favorably associated to QTL alleles. The MAS was applied within family to young bulls, before progeny testing, as part of an open nucleus. Young bulls were selected using strategies based on 1) the single marker with greatest contrast (BEST1), 2) the sum of n greatest contrasts (BESTn), 3) the best n contrasts, limited to one per chromosome (LIMn), 4) the sum of all contrasts exceeding a given threshold n (THRESn), and 5) the sum of contrasts exceeding a threshold, but limited to one per chromosome (LIMT). The maximum progress was achieved by strategies that selected upon several markers flanking multiple QTL in each generation. When THRES was applied, the mean true breeding value (TBV) of selected bulls was increased by 11.98% (over conventional selection) versus 6.73% for BEST1 in the first generation. Applying a full genome scan in each generation allowed selection for different QTL across time. By selecting for multiple QTL over time, MAS maintained superiority over conventional selection for many generations.