Construção de uma biblioteca BAC e avaliação de marcadores para caracterização de regiões alvo do genoma do búfalo

O crescimento da população bubalina em território brasileiro está relacionado ao interesse dos produtores nesse animal, como uma alternativa para a produção de carne, leite e seus derivados. Diante deste panorama, torna-se necessário o aprimoramento de programas de melhoramento genético, visando a seleção de animais geneticamente superiores para a reprodução. Por essa razão, o conhecimento do genoma da espécie é de extrema valia para o setor, uma vez que gera informações necessárias à identificação e avaliação de genes associados com características de interesse econômico. Desse modo, o presente trabalho envolveu a construção de uma nova ferramenta genômica para búfalo, denominada biblioteca BAC, a qual permitirá um novo direcionamento nos estudos moleculares do genoma bubalino, destacando-se a definição da estrutura e organização de genes de interesse econômico, fornecendo informações em nível de seqüência de DNA para o entendimento dos mecanismos e regulação dos mesmos. Com a disponibilidade dessa ferramenta, as primeiras regiões a serem caracterizadas serão aquelas que contêm genes de interesse econômico, as quais se destacam as regiões com genes relacionados com produção e qualidade do leite, regiões com genes relacionados com resposta imune e adaptativa, e regiões com genes da família das lipocalinas, os quais estão envolvidos com características de produção e reprodução. Porém, para que esses genes possam ser caracterizados por meio da biblioteca BAC de búfalo, há a necessidade de gerar marcadores para identificar e isolar clones específicos para esses genes. Marcadores derivados do genoma bovino têm sido utilizados com sucesso em estudos de mapeamento do genoma bubalino, os quais podem ser uma fonte importante de marcadores. Porém, para famílias gênicas, há a necessidade da geração de marcadores específicos para búfalo...The increase of the buffalo population in Brazil is the result of the great interest of the producers as an alternative source for the production of meat, milk and dairy products. Thus, it becomes necessary genetic improvement programs more effective, in order to select genetically superior animals for breeding. The knowledge of the buffalo genome is valuable in this regard, since it generates information to identify and evaluate genes associated with economically traits. In this project we constructed a new genomic tool for buffalo - a genomic BAC library - which can be used in molecular studies of buffalo genome especially those related with the definition of the molecular structure and organization of economically important genes. Once this genomic tool is available, the first target regions of the buffalo genome to be characterized are those containing genes related to milk production, adaptive and innate immune response, and regions lipocalin genes involved in reproduction and production traits. However, to characterize these regions using the BAC library is necessary to have molecular markers to be able to identify and isolate the specific clones. Markers derived from the bovine genome have been successfully used in buffalo genome mapping studies, showing to be an important source of markers. On the another hand, for those genes found in the genome as gene families, there is a need for buffalo specific markers. The evaluation of new markers will contribute to the characterization of those target regions of the buffalo genome, providing information about the genomic architecture of this specie when compared with other bovidFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Construção de um mapa comparativo preliminar do cromossomo 6 do búfalo de rio (Bubalus bubalis) utilizando um painel de células somáticas hídricas irradiadas

Nesse trabalho apresentamos o primeiro mapa comparativo do cromossomo 6 do búfalo de rio (BBU6), desenvolvido a partir da utilização de um painel de células somáticas híbridas irradiadas búfalo-roedor com fragmentação de 5000 rads. O mapa preliminar construído é composto por 33 marcadores, os quais estão ordenados em dois grupos de ligação compostos por 12 microssatélites, 19 genes codificantes e duas ESTs. A freqüência de retenção observada entre os marcadores variou de 14.4% a 40%. A ordem dos marcadores dentro dos grupos de ligação do BBU6, em sua maioria, é consistente com o mapa RH bovino. Esse mapa preliminar do BBU6 é um ponto de partida para comparar a ordem dos genes entre as espécies, fornecendo uma oportunidade para estudar micro-arranjos nesse cromossomo, além de possibilitar a clonagem posicional em búfalo de rio.We present the first radiation hybrid map of BBU6 developed from a recently constructed river buffalo whole-genome radiation hybrid panel (BBURH5000). The preliminary map contains 33 cattle-derived markers, including 12 microsatellites, 19 coding genes and two ESTs, distributed in two linkage groups. The retention frequency of individual markers ranged from 14.4% to 40.0%. Most of the marker order within the linkage groups is consistent with the cattle RH maps. This preliminary BBU6 RH map is the starting point for comparing gene order between both species, presenting opportunity for examination of micro-rearrangements of these chromosomes and, thereby enhancing the possibility of positional candidate cloning in river buffalo.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Detection of Genes Related to Growth Traits in Sheep in Copy Number Variation Regions

            In Brazil, the Santa Ines sheep breed has aptitude for meat production, standing out for its rusticity and high productivity. Genomic studies in poorly studied breeds are of great importance, as they make it possible, through genotyping, to find sources of structural variations in the sheep genome. Copy number variations (CNV) are the major source of structural variation among individuals, which are defined as modifications in the genome where the number of copies of a genomic DNA sequence differs from a reference genome. These studies make it possible to advance knowledge of the genomes of poorly studied species such as sheep. Thus, the aim of this study was to identify genes related to growth traits in Santa Ines sheep bred in copy number variation regions (CNVR). A total of 638 animals were genotyped with the Ovine SNP50 Genotyping BeadChip (Ilumina). Markers with unknown genomic position and located on sex chromosomes in the reference genome Oar_v 3.1 were excluded. The adjustment of the GC content was performed to reduce the rate of false positive CNVs, considering 500 Kb around each SNP. CNV detection was performed with the PennCNV software, where CNVs identified in only one sample, with fewer than three SNPs, BAF drift>0.01 and GC wave factor>0.09 were excluded. CNVRs were inferred by concatenating individual CNVs identified in more than one animal using the CNVRuler software, excluding genomic regions with an allelic frequency lower than 0.5% and with recurrence <0.1. A total of 742 CNVs (357 deletions and 385 duplications) were detected, which were grouped in 360 CNVRs (224 gain, 132 loss and 4 both) detected on all autosomal chromosomes. The CNVRs inferred in this study had an average size of 142,655 bp (from 12,139 to 2,643,483 bp), covered 2.03% of the autosomal genome sequence, and had frequencies ranging from 0.31% to 4.85% in this population. The OAR1 showed the largest number of CNVRs (40), while the OAR20 presented the smallest number of CNVRs (3). A total of 832 Ensembl ID from protein-coding genes were identified in these CNVRs. Subsequently, functional enrichment analysis was performed with the DAVID software and revealed 44 Gene Ontology terms and 8 enriched KEGG pathways (P<0.05). Among the genes identified in the CNVRs, we highlight the following genes related to muscle growth and development: CCNT2, UNC45B, DYNC1LI2, KLHL40, MYH1, CORO1B and KMT5B. In addition, we also identified genes related to ossification, such as NLE1, W5P5S7_SHEEP, COL1A1, TCIRG1, GIT1, W5P1A2_SHEEP and BCL. The results of the present study provide information to help understand the genetic architecture of growth traits in sheep, their possible impacts on productive traits, and future investigations, being especially useful for researchers interested in structural variations in the sheep genome

A genome-wide single nucleotide polymorphism and copy number variation analysis for number of piglets born alive

Abstract Background In this study we integrated the CNV (copy number variation) and WssGWAS (weighted single-step approach for genome-wide association) analyses to increase the knowledge about number of piglets born alive, an economically important reproductive trait with significant impact on production efficiency of pigs. Results A total of 3892 samples were genotyped with the Porcine SNP80 BeadChip. After quality control, a total of 57,962 high-quality SNPs from 3520 Duroc pigs were retained. The PennCNV algorithm identified 46,118 CNVs, which were aggregated by overlapping in 425 CNV regions (CNVRs) ranging from 2.5 Kb to 9718.4 Kb and covering 197 Mb (~ 7.01%) of the pig autosomal genome. The WssGWAS identified 16 genomic regions explaining more than 1% of the additive genetic variance for number of piglets born alive. The overlap between CNVR and WssGWAS analyses identified common regions on SSC2 (4.2–5.2 Mb), SSC3 (3.9–4.9 Mb), SSC12 (56.6–57.6 Mb), and SSC17 (17.3–18.3 Mb). Those regions are known for harboring important causative variants for pig reproductive traits based on their crucial functions in fertilization, development of gametes and embryos. Functional analysis by the Panther software identified 13 gene ontology biological processes significantly represented in this study such as reproduction, developmental process, cellular component organization or biogenesis, and immune system process, which plays relevant roles in swine reproductive traits. Conclusion Our research helps to improve the understanding of the genetic architecture of number of piglets born alive, given that the combination of GWAS and CNV analyses allows for a more efficient identification of the genomic regions and biological processes associated with this trait in Duroc pigs. Pig breeding programs could potentially benefit from a more accurate discovery of important genomic regions

Genetic parameters, genetic trends, and principal component analysis for productive and reproductive traits of Guzera beef cattle

The objectives of this study were to estimate genetic parameters and genetic trends and perform principal component analysis for the following traits: body weight adjusted at 210 days of age (W210), one year (W365), and yearling or 450 days of age (W450); weight gain from birth to weaning or 210 days of age (WGBW), from weaning to 365 days of age (WGW365), and from one year to yearling (WGY); and scrotal circumference adjusted at 365 (SC365) and 450 (SC450) days of age of Guzera beef cattle. Variance components were estimated under a two-trait animal model using the restricted maximum likelihood method. The fixed effects included in the model were birth season, contemporary group, and covariable age of dam at calving as linear and quadratic effects. Heritability estimates ranged from 0.10 (WGY) to 0.41 (W450 and SC450). The studied traits could be used as selection criteria because of their sufficient additive genetic variability to respond to the selection process. However, lower selection efficiency is expected for WGBW and WGY. Genetic trends studied were significant for all traits, except WGY. The W365 and W450 traits belonged to the “Genetic Merit Total” index and responded to selection with favorable genetic gain per year. The traits that were not included in the selection index showed favorable positive genetic correlations attributed to the indirect selection processes. The principal component analysis identified two groups of traits. The first group related to body weight and weight gain and the second related to the scrotal circumference, indicating that selection could be applied to one group of traits within each observed group. The genetic gains were attributed in the same direction for the other traits belonging to the groups. Thus, we recommend selecting animals for W365 based on first group, and SC365 based on secon

Association of apolipoprotein B and adiponectin receptor 1 genes with carcass, bone integrity and performance traits in a paternal broiler line

Apolipoprotein B (APOB) and Adiponectin Receptor 1 (ADIPOR1) are related to the regulation of feed intake, fat metabolism and protein deposition and are candidate genes for genomic studies in birds. In this study, associations of two single nucleotide polymorphisms (SNPs) g.102A>T (APOB) and g.729C>T (ADIPOR1) with carcass, bone integrity and performance traits in broilers were investigated. Genotyping was performed on a paternal line of 1,454 broilers. The SNP detection was carried out by PCR-RFLP technique using the restriction enzymes HhaI for the SNP g.729C>T and MslI for the SNP g.102A>T. The association analyses of the two SNPs with 85 traits were performed using the restricted maximum likelihood (REML) and Generalized Quasi-Likelihood Score (GQLS) methods. For REML the model included the random additive genetic effect of animal and fixed effects of sex, hatch and SNP genotypes. In the GQLS method, a logistic regression was used to associate the genotypes with phenotypes adjusted for fixed effects of sex and hatch. The SNP g.729C>T in the ADIPOR1 gene was associated with thickness of the femur and breast skin yield. Thus, the ADIPOR1 gene seems implicated in the metabolism and/or fat deposition and bone integrity in broilers.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Genomic regions and pathways associated with gastrointestinal parasites resistance in Santa Inês breed adapted to tropical climate

Abstract Background The aim of this study was to estimate variance components and to identify genomic regions and pathways associated with resistance to gastrointestinal parasites, particularly Haemonchus contortus, in a breed of sheep adapted to tropical climate. Phenotypes evaluations were performed to verify resistance to gastrointestinal parasites, and were divided into two categories: i) farm phenotypes, assessing body condition score (BCS), degree of anemia assessed by the famacha chart (FAM), fur score (FS) and feces consistency (FC); and ii) lab phenotypes, comprising blood analyses for hematocrit (HCT), white blood cell count (WBC), red blood cell count (RBC), hemoglobin (HGB), platelets (PLT) and transformed (log10) egg per gram of feces (EPGlog). A total of 576 animals were genotyped with the Ovine SNP12k BeadChip (Illumina, Inc.), that contains 12,785 bialleleic SNP markers. The variance components were estimated using a single trait model by single step genomic BLUP procedure. Results The overall linkage disequilibrium (LD) mean between pairs of markers measured by r 2 was 0.23. The overall LD mean between markers considering windows up to 10 Mb was 0.07. The mean LD between adjacent SNPs across autosomes ranged from 0.02 to 0.10. Heritability estimates were low for EPGlog (0.11), moderate for RBC (0.18), PLT (0.17) HCT (0.20), HGB (0.16) and WBC (0.22), and high for FAM (0.35). A total of 22, 21, 23, 20, 26, 25 and 23 windows for EPGlog for FAM, WBC, RBC, PLT, HCT and HGB traits were identified, respectively. Among the associated windows, 10 were shown to be common to HCT and HGB traits on OAR1, OAR2, OAR3, OAR5, OAR8 and OAR15. Conclusion The traits indicating gastrointestinal parasites resistance presented an adequate genetic variability to respond to selection in Santa Inês breed, and it is expected a higher genetic gain for FAM trait when compared to the others. The level of LD estimated for markers separated by less than 1 Mb indicated that the Ovine SNP12k BeadChip might be a suitable tool for identifying genomic regions associated with traits related to gastrointestinal parasite resistance. Several candidate genes related to immune system development and activation, inflammatory response, regulation of lymphocytes and leukocytes proliferation were found. These genes may help in the selection of animals with higher resistance to parasites