7 research outputs found
Secuenciaci贸n de mol茅culas individuales de 谩cidos nucleicos en tiempo real (SMRT) para caracterizar transcriptomas e isoformas de ARNm
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.Nuestros esfuerzos est谩n orientados a evaluar patrones de expresi贸n g茅nica del cromosoma Y bovino. Los genes de inter茅s son los denominados genes X-degenerados que se encuentran en la regi贸n espec铆fica masculina del cromosoma Y (MSY). Esta regi贸n contiene el 95% del ADN del cromosoma Y. Estos genes son de copia 煤nica y tienen un hom贸logo en el cromosoma X. Ambos hom贸logos tienen perfiles amplios de expresi贸n. Sin embargo, algunos genes, como el SRY que regula la determinaci贸n del sexo masculino, tienen funciones m谩s espec铆ficas. La identificaci贸n de las diferencias de secuencia de ADN entre estos hom贸logos permitir谩 evaluar sus patrones de expresi贸n espacial y temporal. La identificaci贸n de los ARNm codificados en el cromosoma Y y de sus isoformas permitir谩n analizar la expresi贸n espec铆fica de sus isoformas en tejidos masculinos. Esto 煤ltimo facilitar谩 nuestra evaluaci贸n de funci贸n g茅nica en la diferenciaci贸n sexual masculina y la fertilidad. Por lo tanto, planteamos la hip贸tesis de que cada uno de estos genes hom贸logos degenerados del X genera isoformas y que existen patrones de expresi贸n diferencial entre sexos y tejidos. Para investigar esto 煤ltimo, utilizamos una tecnolog铆a de secuenciaci贸n de nueva generaci贸n (NGS) que genera lecturas de secuenciaci贸n largas con un rango de longitud de 1000 a 10,000 pares de bases. Se secuenciaron los transcriptomas en varios tejidos (h铆gado, pulm贸n, adiposo, muscular, hipot谩lamo y test铆culo). Se utilizaron las secuencias generadas para el an谩lisis bioinform谩tico y la caracterizaci贸n de isoformas. Siendo el foco de este manuscrito la tecnolog铆a SMRT, solo presentamos los resultados obtenidos con el an谩lisis de los genes bUTY y bUTX
Cytogenetic Mapping of 35 New Markers in the Alpaca (Vicugna pacos)
Universidad Nacional Agraria La Molina. Escuela de Posgrado. Maestr铆a en Producci贸n AnimalAlpaca 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鈥揷amelid 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. View Full-Tex
Development of a bovine X chromosome linkage group and painting probes to assess cattle, sheep, and goat X chromosome segment homologies
The X chromosome linkage group is conserved in placental mammals. However, X chromosome morphological differences, due to internal chromosome rearrangements, exist among mammalian species. We have developed bovine chromosome painting probes for Xp and Xq to assess segment homologies between the submetacentric bovine X chromosome and the acrocentric sheep and goat X chromosomes. These painting probes and their corresponding DNA libraries were developed by chromosome micromanipulation, DNA micropurification, microcloning, and PCR amplification. The bovine Xp painting probe identified an interstitially located homologous segment in the sheep and goat Xq region, most probably resulting from chromosome inversion. Ten type II (microsatellite) markers obtained from the bovine Xq library and five other X chromosome assigned, but unlinked, markers were used to generate a linkage map for Xq spanning 89.4 centimorgans. The chromosome painting probes and molecular markers generated in this study would be useful for comparative mapping and tracing of internal X chromosome rearrangements in all ruminant species and would contribute to the understanding of mammalian sex chromosome evolution
Genome-Wide Association Study of Fiber Diameter in Alpacas
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.Ministerio de Desarrollo Agrario y Riego (Per煤)Universidad Nacional Agraria La Molina (Per煤)Depto. de Producci贸n AnimalFac. de VeterinariaTRUEpubDescuento UC