RESUMO
The aim of this study was the identification of candidate genomic regions associated with fiber diameter in alpacas. DNA samples were collected from 1011 female Huacaya alpacas from two geographical Andean regions in Peru (Pasco and Puno), and three alpaca farms within each region. The samples were genotyped using an Affymetrix Custom Alpaca genotyping array containing 76,508 SNPs. After the quality controls, 960 samples and 51,742 SNPs were retained. Three association study methodologies were performed. The GWAS based on a linear model allowed us to identify 11 and 35 SNPs (-log10(p-values) > 4) using information on all alpacas and alpacas with extreme values of fiber diameter, respectively. The haplotype and marker analysis method allowed us to identify nine haplotypes with standardized haplotype heritability higher than six standard deviations. The selection signatures based on cross-population extended haplotype homozygosity (XP-EHH) allowed us to identify 180 SNPs with XP-EHH values greater than |3|. Four candidate regions with adjacent SNPs identified via two association methods of analysis are located on VPA6, VPA9, VPA29 and one chromosomally unassigned scaffold. This study represents the first analysis of alpaca whole genome association with fiber diameter, using a recently assembled alpaca SNP microarray.
RESUMO
Alpaca is a camelid species of broad economic, biological and biomedical interest, and an essential part of the cultural and historical heritage of Peru. Recently, efforts have been made to improve knowledge of the alpaca genome, and its genetics and cytogenetics, to develop molecular tools for selection and breeding. Here, we report cytogenetic mapping of 35 new markers to 19 alpaca autosomes and the X chromosome. Twenty-eight markers represent alpaca SNPs, of which 17 are located inside or near protein-coding genes, two are in ncRNA genes and nine are intergenic. The remaining seven markers correspond to candidate genes for fiber characteristics (BMP4, COL1A2, GLI1, SFRP4), coat color (TYR) and development (CHD7, PAX7). The results take the tally of cytogenetically mapped markers in alpaca to 281, covering all 36 autosomes and the sex chromosomes. The new map assignments overall agree with human-camelid conserved synteny data, except for mapping BMP4 to VPA3, suggesting a hitherto unknown homology with HSA14. The findings validate, refine and correct the current alpaca assembly VicPac3.1 by anchoring unassigned sequence scaffolds, and ordering and orienting assigned scaffolds. The study contributes to the improvement in the alpaca reference genome and advances camelid molecular cytogenetics.
Assuntos
Camelídeos Americanos/genética , Mapeamento Cromossômico/veterinária , Animais , Células Cultivadas , Cromossomos Artificiais Bacterianos , Análise Citogenética , Marcadores Genéticos , Genoma , Hibridização in Situ Fluorescente , Polimorfismo de Nucleotídeo Único , Cromossomos Sexuais/genética , Fibra de LãRESUMO
Abstract Developing countries have the challenge of achieving food security in a world context that is affected by climate change and global population growth. Molecular Genetics and genomics are proposed as technologies that will help to achieve sustainable food security. Technologies that have been developed in the last decade such as the development of genetic markers, genetic maps, genomic selection, next-generation sequencing, and DNA editing systems are discussed. Examples of some discoveries and achievements are provided.
Resumen Los países en vías de desarrollo tienen el reto de alcanzar seguridad alimentaria en un contexto mundial afectado por el cambio climático y crecimiento poblacional global. La genética molecular y la genómica son propuestas como tecnologías que ayudarán a alzanzar una seguridad alimentaria sostenible. Tecnologías que han sido desarrolladas en la última década como el desarrollo de marcadores moleculares, mapeo genético, selección genómica, secuenciamiento de próxima generación y sistemas de edición de ADN son discutidos. Se proveen ejemplos de algunos descubrimientos y logros.
RESUMO
Abstract Our efforts are oriented to assess bovine Y-chromosome gene expression patterns. One set of genes that are of interest are the so-called X-degenerate Y-chromosome genes that are located in the male-specific region of the Y-chromosome (MSY). This region contains 95% of the DNA of the Y chromosome. These genes are single copy and have an X-chromosome homolog. Both, the Y-encoded and X-encoded homologs have ubiquitous expression profiles. However, some genes, like SRY that regulates male sex determination, have functions that are more specific. Identifying DNA sequence differences between these homologs will allow evaluation of their spatial and temporal expression patterns. Identification of the Y-encoded mRNAs and their isoforms will allow our understanding of tissue specific expression of isoforms in male tissues. The latter will facilitate our evaluation of gene function in male sex differentiation and fertility. Hence, we hypothesized that each of these X-degenerate gene homologs generate isoforms and that differential expression patterns exist between sexes and across tissues. To investigate the latter we used a new generation sequencing (NGS) technology that generates long sequencing reads with a range between 1000 to 10,000 base pairs in length. Single molecule real time (SMRT) isoform sequencing (IsoSeq) of several tissues (liver, lung, adipose, muscle, hypothalamus and testis) was carried out. Transcript sequences were used for bioinformatics analysis and isoform characterization. Given the focus of this manuscript the SMRT technology we are only presenting results obtained with the analysis of the bUTY and bUTX genes.
Resumen Nuestros esfuerzos están orientados a evaluar patrones de expresión génica del cromosoma Y bovino. Los genes de interés son los denominados genes X-degenerados que se encuentran en la región específica masculina del cromosoma Y (MSY). Esta región contiene el 95% del ADN del cromosoma Y. Estos genes son de copia única y tienen un homólogo en el cromosoma X. Ambos homólogos tienen perfiles amplios de expresión. Sin embargo, algunos genes, como el SRY que regula la determinación del sexo masculino, tienen funciones más específicas. La identificación de las diferencias de secuencia de ADN entre estos homólogos permitirá evaluar sus patrones de expresión espacial y temporal. La identificación de los ARNm codificados en el cromosoma Y y de sus isoformas permitirán analizar la expresión específica de sus isoformas en tejidos masculinos. Esto último facilitará nuestra evaluación de función génica en la diferenciación sexual masculina y la fertilidad. Por lo tanto, planteamos la hipótesis de que cada uno de estos genes homólogos degenerados del X genera isoformas y que existen patrones de expresión diferencial entre sexos y tejidos. Para investigar esto último, utilizamos una tecnología de secuenciación de nueva generación (NGS) que genera lecturas de secuenciación largas con un rango de longitud de 1000 a 10,000 pares de bases. Se secuenciaron los transcriptomas en varios tejidos (hígado, pulmón, adiposo, muscular, hipotálamo y testículo). Se utilizaron las secuencias generadas para el análisis bioinformático y la caracterización de isoformas. Siendo el foco de este manuscrito la tecnología SMRT, solo presentamos los resultados obtenidos con el análisis de los genes bUTY y bUTX.
RESUMO
The alpaca (Vicugna pacos) is an economically important and cultural signature species in Peru. Thus, molecular genomic information about the genes underlying the traits of interest, such as fiber properties and color, is critical for improved breeding and management schemes. Current knowledge about the alpaca genome, particularly the chromosomal location of such genes of interest is limited and lags far behind other livestock species. The main objective of this work was to localize alpaca candidate genes for fiber growth and color using fluorescence in situ hybridization (FISH). We report the mapping of candidate genes for fiber growth COL1A1, CTNNB1, DAB2IP, KRT15, KRTAP13-1, and TNFSF12 to chromosomes 16, 17, 4, 16, 1, and 16, respectively. Likewise, we report the mapping of candidate genes for fiber color ALX3, NCOA6, SOX9, ZIC1, and ZIC5 to chromosomes 9, 19, 16, 1, and 14, respectively. In addition, since KRT15 clusters with five other keratin genes (KRT31, KRT13, KRT9, KRT14, and KRT16) in scaffold 450 (Vic.Pac 2.0.2), the entire gene cluster was assigned to chromosome 16. Similarly, mapping NCOA6 to chromosome 19, anchored scaffold 34 with 8 genes, viz., RALY, EIF2S2, XPOTP1, ASIP, AHCY, ITCH, PIGU, and GGT7 to chromosome 19. These results are concordant with known conserved synteny blocks between camelids and humans, cattle and pigs.
RESUMO
Alpacas are one of four South American Camelid species living in the highlands of the Andes. Production of alpaca fiber contributes to the economy of the region and the livelihood of many rural families. Fiber quantity and quality are important and in need of a modern breeding program based on genomic selection to accelerate genetic gain. To achieve this is necessary to discover enough molecular markers, single nucleotide polymorphisms (SNPs) in particular, to provide genome coverage and facilitate genome wide association studies to fiber production characteristics. The aim of this study was to discover alpaca SNPs by genotyping forty alpaca DNA samples using the BovineHD Genotyping Beadchip. Data analysis was performed with GenomeStudio (Illumina) software. Because different filters and thresholds are reported in the literature we investigated the effects of no-call threshold (≥0.05, ≥0.15, and ≥0.25) and call frequency (≥0.9 and =1.0) in identifying positive SNPs. Average GC Scores, calculated as the average of the 10% and 50% GenCall scores for each SNP (≥0.70) and the GenTrain score ≥ 0.25 parameters were applied to all comparisons. SNPs with minor allele frequency (MAF) ≥ 0.05 or ≥ 0.01 were retained. Since detection of SNPs is based on the stable binding of oligonucleotide probes to the target DNA immediately adjacent to the variant nucleotide, all positive SNP flanking sequences showing perfect alignments between the bovine and alpaca genomes for the first 21 or 26 nucleotides flanking the variant nucleotide at either side were selected. Only SNPs localized in one scaffold were assumed unique. Unique SNPs identified in both reference genomes were kept and mapped on the Vicugna_pacos 2.0.2 genome. The effects of the no-call threshold ≥ 0.25, call frequency = 1 and average GC ≥ 0.7 were meaningful and identified 6756 SNPs of which 400 were unique and polymorphic (MAF ≥ 0.01). Assignment to alpaca chromosomes was possible for 292 SNPs. Likewise, 209 SNPs were localized in 202 alpaca gene loci and 29 of these share the same loci with the dromedary. Interestingly, 69 of 400 alpaca SNPs have 100% similarity with dromedary.
RESUMO
The decrease in sequencing cost and increased sophistication of assembly algorithms for short-read platforms has resulted in a sharp increase in the number of species with genome assemblies. However, these assemblies are highly fragmented, with many gaps, ambiguities, and errors, impeding downstream applications. We demonstrate current state of the art for de novo assembly using the domestic goat (Capra hircus) based on long reads for contig formation, short reads for consensus validation, and scaffolding by optical and chromatin interaction mapping. These combined technologies produced what is, to our knowledge, the most continuous de novo mammalian assembly to date, with chromosome-length scaffolds and only 649 gaps. Our assembly represents a â¼400-fold improvement in continuity due to properly assembled gaps, compared to the previously published C. hircus assembly, and better resolves repetitive structures longer than 1 kb, representing the largest repeat family and immune gene complex yet produced for an individual of a ruminant species.
Assuntos
Cromatina/genética , Genoma/genética , Cabras/genética , Animais , Cromossomos/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequências Repetitivas de Ácido Nucleico/genéticaRESUMO
Polydactyly is a common malformation of vertebrate limbs. Preaxial polydactyly (PPD) has been mapped in human, mouse and chicken to the syntenic region of human 7q36. Lmbr1 was thought as the critical candidate gene for human and mouse PPD. To understand the molecular mechanism underlying chicken polydactyly, we have cloned the open reading frame (ORF) of chicken Lmbr1, which contains 1467 nucleotides. Within this ORF, we found one short and one long splice forms. The short splice form has a complete deletion of exon 4. Six cSNPs were found in the chicken ORF, and two of these cSNPs, G797A and G1255A, lead to amino acid substitutions. However, G797A substitution had no significant association with polydactyly and the G1255A substitution had very low frequency in the population. The T1254C polymorphism in exon 13 was found to be strongly associated with polydactyly. Radiation hybrid mapping of a DNA fragment containing intron 13 of the chicken Lmbr1 assigned the gene to chromosome 2 between MCW071 (a marker within the EN2 gene) and ADL0270, a syntenic region to human 7q36.
Assuntos
Galinhas/genética , Polidactilia/genética , Polimorfismo de Nucleotídeo Único , Alelos , Processamento Alternativo , Substituição de Aminoácidos , Animais , Sequência de Bases , Cromossomos , Clonagem Molecular , Éxons , Ligação Genética , Íntrons , Proteínas de Membrana/genética , Dados de Sequência Molecular , Fases de Leitura Aberta , Polidactilia/etiologia , Polimorfismo Conformacional de Fita Simples , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Mapeamento de Híbridos RadioativosRESUMO
Characterization of the Gallus gallus alpha-amylase gene family revealed that the chicken genome contains two distinct amy loci. One of the two loci is expressed in the chicken pancreas while cDNA clones for the second locus were detected in a library constructed from liver mRNA. Fluorescent in situ hybridization to chromosome spreads showed that the two loci are both located on chromosome 8 within the chicken genome. Moreover, each locus contains both an intact, expressed gene copy as well as a pseudogene. The expressed gene and the pseudogene are arranged in a divergent configuration in the pancreatic amy locus, while in the hepatic locus the intact gene and the pseudogene are arranged in tandem. The data suggest a complex pattern of evolution for the chicken amylase gene family which includes multiple gene duplication events, insertion/deletion events, as well as changes in spatial expression patterns.
Assuntos
Galinhas/genética , alfa-Amilases/genética , Sequência de Aminoácidos , Animais , Mapeamento Cromossômico , Códon de Terminação , Dosagem de Genes , Fígado/metabolismo , Modelos Biológicos , Dados de Sequência Molecular , Pâncreas/metabolismo , Pseudogenes , Homologia de Sequência de AminoácidosRESUMO
Screening a bovine Y Chromosome-specific DNA library resulted in 34 new microsatellites, six of which mapped to the pseudoautosomal region (PAR), and 28 localized to the Y-specific region. These microsatellites, together with 23 markers previously mapped to the bovine Y Chr, were scored on a 7000-rad cattle-hamster radiation hybrid (RH) panel. Retention frequency of individual markers ranged from 18.5% to 76.5% with an average of 48.4%. Markers with high retention frequency (>55%) were found to exist in multiple copies on the Y Chr. Thirteen markers were placed on the PAR RH map with the AmelY gene proximal to the pseudoautosomal boundary and 46 markers, including Sry and Tspy gene, on the Y-specific region of the RH map. The microsatellites developed and mapped in this work will be useful for comparative mapping of cattle, sheep, and goat, studying the origin, evolution, and migration of bovidae species and provide an initial platform to develop a high-resolution map of the Y Chr and positional cloning of Y-specific genes.
