RESUMO
Wool is produced via synthetic processes of wool follicles, which are embedded in the skin of sheep. The development of new-generation sequencing and RNA sequencing provides new approaches that may elucidate the molecular regulation mechanism of wool follicle development and facilitate enhanced selection for wool traits through gene-assisted selection or targeted gene manipulation. We performed de novo transcriptome sequencing of skin using the Illumina Hiseq 2000 sequencing system in sheep (Ovis aries). Transcriptome de novo assembly was carried out via short-read assembly programs, including SOAPdenovo and ESTScan. The protein function, clusters of orthologous group function, gene ontology function, metabolic pathway analysis, and protein coding region prediction of unigenes were annotated by BLASTx, BLAST2GO, and ESTScan. More than 26,266,670 clean reads were collected and assembled into 79,741 unigene sequences, with a final assembly length of 35,447,962 nucleotides. A total of 22,164 unigenes were annotated, accounting for 36.27% of the total number of unigenes, which were divided into 25 classes belonging to 218 signaling pathways. Among them, there were 17 signal paths related to hair follicle development. Based on mass sequencing data of sheepskin obtained by RNA-Seq, many unigenes were identified and annotated, which provides an excellent platform for future sheep genetic and functional genomic research. The data could be used for improving wool quality and as a model for human hair follicle development or disease prevention.
Assuntos
Carneiro Doméstico/genética , Pele/metabolismo , Transcriptoma , Lã/fisiologia , Animais , Mapeamento de Sequências Contíguas , Etiquetas de Sequências Expressas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Biblioteca Gênica , Genômica , Modelos Genéticos , Anotação de Sequência Molecular , Análise de Sequência de RNARESUMO
This study investigated geographic and pairwise distances among seven Chinese local and four introduced sheep populations via analysis of 26 microsatellite DNA markers. Genetic polymorphism was rich, and the following was discovered: 348 alleles in total were detected, the average allele number was 13.38, the polymorphism information content (PIC) of loci ranged from 0.717 to 0.788, the number of effective alleles ranged from 7.046 to 7.489, and the observed heterozygosity ranged from 0.700 to 0.768 for the practical sample, and from 0.712 to 0.794 for expected heterozygosity. The Wright's F-statistic of subpopulations within the total (FST) was 0.128, the genetic differentiation coefficient (GST) was 0.115, and the average gene flow (Nm) was 1.703. The phylogenetic trees based on the neighbor-joining method by Nei's genetic distance (DA) and Nei's standard genetic distance (DS) were similar. Sheep populations clustered into group 1 (Ta, M, L, H, O, G, and Q breeds) and group 2 (PD, WS, B, and T breeds). These results will have an important value applied and directive significance for sheep breeding in the future.
Assuntos
Repetições de Microssatélites , Ovinos/genética , Animais , Evolução Molecular , Deriva Genética , Marcadores Genéticos , Variação Genética , Espécies Introduzidas , Filogeografia , Análise de Sequência de DNA , Ovinos/sangueRESUMO
Variation in microsatellite or simple sequence repeat (SSR) loci has, until recently, relied heavily on the use of gel-based methods that can be both time consuming and difficult to genotype. Non gel-based systems are therefore important to increase simplicity and improve turn-around time without compromising assay sensitivity and accuracy. In this report, we assessed the latest of the non-gel-based methods, high-resolution melting (HRM) curve analysis. HRM is a technique that monitors exactly the decreasing fluorescence of intercalating dye in the process of dissociation of double-stranded DNA. The measurement immediately follows polymerase chain reaction in a one-step, closed-tube method. Four SSR loci of different complexity in sheep, namely MAF209, MCM140, CB226, and SRCRSP5, were assessed using the LightScanners System with LC Greens PLUS DNA binding dye. In order to improve the accuracy of genotyping, we applied internal oligo nucleotide calibrators while performing HRM. DNA polymorphisms were previously identified using capillary electrophoresis analysis (CE). The result showed that CE detected more genotypes than HRM in the same loci regardless of the level of polymorphism at the SSR loci. We demonstrate current limitations of the HRM method for the analysis of SSR loci.