MicroRNAs (miRNAs) are a class of small RNAs that play significant roles in regulating the expression of the post-transcriptional skin and hair follicle gene. In recent years, extensive studies on these microRNAs have been carried out in mammals such as mice, rats, pigs and cattle. By comparison, the number of microRNAs that have been identified in goats is relatively low; and in particular, the miRNAs associated with the processes of skin and hair follicle development remain largely unknown. In this study, areas of skin where the cashmere grows in anagen were sampled. A total of 10,943,292 reads were obtained using Solexa sequencing, a high-throughput sequencing technology. From 10,644,467 reads, we identified 3,381 distinct reads and after applying the classification statistics we obtained 316 miRNAs. Among them, using conservative identification, we found that 68 miRNAs (55 of these are confirmed to match known sheep and goat miRNAs in miRBase ) are conserved in goat and have been reported in NCBI; the remaining 248 miRNA were conserved in other species but have not been reported in goat. Furthermore, we identified 22 novel miRNAs. Both the known and novel miRNAs were confirmed by a second sequencing using the same method as was used in the first. This study confirmed the authenticity of 316 known miRNAs and the discovery of 22 novel miRNAs in goat. We found that the miRNAs that were co-expressed in goat and sheep were located in the same region of the respective chromosomes and may play an essential role in skin and follicle development. Identificaton of novel miRNAs resulted in significant enrichment of the repertoire of goat miRNAs.

Inner Mongolia and Liaoning cashmere goats are two outstanding Chinese multipurpose breeds that adapt well to the semi-arid temperate grassland. These two breeds are characterized by their soft cashmere fibers, thus making them great models to identify genomic regions that are associated with cashmere fiber traits. Whole-genome sequencing of 70 cashmere goats produced more than 5.52 million single-nucleotide polymorphisms and 710,600 short insertions and deletions. Further analysis of these genetic variants showed some population-specific molecular markers for the two cashmere goat breeds that are otherwise phenotypically similar. By analyzing F_ST and θ_π outlier values, we identified 135 genomic regions that were associated with cashmere fiber traits within the cashmere goat populations. These selected genomic regions contained genes, which are potential involved in the production of cashmere fiber, such as FGF5, SGK3, IGFBP7, OXTR, and ROCK1. Gene ontology enrichment analysis of identified short insertions and deletions also showed enrichment in keratinocyte differentiation and epidermal cell differentiation. These findings demonstrate that this genomic resource will facilitate the breeding of cashmere goat and other Capra species in future.

It is broadly expected that next generation sequencing will ultimately generate a complete genome as is the latest goat reference genome (ARS1), which is considered to be one of the most continuous assemblies in livestock. However, the rich diversity of worldwide goat breeds indicates that a genome from one individual would be insufficient to represent the whole genomic contents of goats. By comparing nine de novo assemblies from seven sibling species of domestic goat with ARS1 and using resequencing and transcriptome data from goats for verification, we identified a total of 38.3 Mb sequences that were absent in ARS1. The pan-sequences contain genic fractions with considerable expression. Using the pan-genome (ARS1 together with the pan-sequences) as a reference genome, variation calling efficacy can be appreciably improved. A total of 56,657 spurious SNPs per individual were repressed and 24,414 novel SNPs per individual on average were recovered as a result of better reads mapping quality. The transcriptomic mapping rate was also increased by ∼1.15%. Our study demonstrated that comparing de novo assemblies from closely related species is an efficient and reliable strategy for finding missing sequences from the reference genome and could be applicable to other species. Pan-genome can serve as an improved reference genome in animals for a better exploration of the underlying genomic variations and could increase the probability of finding genotype-phenotype associations assessed by a comprehensive variation database containing much more differences between individuals. We have constructed a goat pan-genome web interface for data visualization (http://animal.nwsuaf.edu.cn/panGoat).

Crossovers generated by homologous recombination ensure proper chromosome segregation during meiosis. Crossover interference results in chiasmata being more evenly distributed along chromosomes, but the mechanism underlying crossover interference remains elusive. Based on large pedigrees of Holstein and Jersey cattle with genotype data, we extracted three-generation families, including 147,327 male and 71,687 female meioses in Holstein, and 108,163 male and 37,008 female meioses in Jersey, respectively. We identified crossovers in these meioses and fitted the Housworth-Stahl “interference-escape” model to study crossover interference patterns in the cattle genome. Our result reveals that the degree of crossover interference is stronger in females than in males. We found evidence for inter-chromosomal variation in the level of crossover interference, with smaller chromosomes exhibiting stronger interference. In addition, crossover interference levels decreased with maternal age. Finally, sex-specific GWAS analyses identified one locus near the NEK9 gene on chromosome 10 to have a significant effect on crossover interference levels. This locus has been previously associated with recombination rate in cattle. Collectively, this large-scale analysis provided a comprehensive description of crossover interference across chromosome, sex and age groups, identified associated candidate genes, and produced useful insights into the mechanism of crossover interference.

Inner Mongolia cashmere goats, as an important part of animal husbandry production, play an important role in animal fiber industry. In recent years, scientific research has made a lot of explorations on the molecular regulation mechanism of hair follicle cycle growth, but few studies have been reported on the development of cashmere hair in fetal period. This study was based on the completion of 21 skin samples of mRNA and miRNA sequencing in 7 fetal periods (45 days, 55 days,65 days,75 days,95 days,115 days and 135 days) of the Inner Mongolia Cashmere goat. The target genes of miRNA associated with the development of secondary hair follicles in the cashmere goats were selected through the combination analysis of mRNA and miRNA data. Then the overexpression vector was constructed and the interaction between the miRNA and the target gene was identified by Dual-Luciferase Reporter Gene System. The function and interaction relationship of chi-miR-199a-5p and TGF-β2 were verified by RT-qPCR and western blot at the level of the fibroblasts in Inner Mongolia Cashmere goat. It provides a theoretical basis for further study of miRNA and its target genes regulating the occurrence and development of skin hair follicles. As the result shows, the expression trends of 7 genes (BAMBI, SMAD1, LTBP1, PPP2R1B, ID4, BMP8B and PITX2) and 7 miRNA (chi-miR-17-5p, chi-miR-125b-3p, chi-miR-21-5p, chi-miR-143-5p and chi-miR-106b-5p) in the skin samples for the seven stages of the fetus were shown to be consistent with the sequencing results. the results of sequencing are reliable. The correlation coefficient of TGF-β2 and chi-miR-199a-5p in fetal 45d-135d expression is −0.84, showing a strong negative correlation, The target relationship was preliminarily judged. The results of double luciferase vector report showed that chi-miR-199a-5p significantly decreased the expression of luciferase in TGF-β2 3′UTR, It is determined that there is a reciprocal relationship between them at a specific time. We transfected chi-miR199a-5p-FAM mimics into fibroblasts cultured in vitro from Inner Mongolia cashmere goats. After transfection, the cells were harvested to extract total RNA and protein. The mRNA and protein expression levels of TGF-β2 in fibroblasts were detected by RT-qPCR and western blot. It was verified that chi-miR-199a-5p inhibited TGF-β2 expression at both mRNA and protein translation levels in fibroblasts. At the same time, it was again proved that the TGF-β2 gene is a target gene of chi-miR199a-5p.