Jinquan Li

Identification of Conserved and Novel microRNAs in Cashmere Goat Skin by Deep Sequencing

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.

Characterization of BMP2 gene expression in embryonic and adult Inner Mongolia Cashmere goat (Capra hircus) hair follicles

Mammalian skin consists of thousands of hair follicles, each undergoing continuous regenerative cycling including anagen, catagen and telogen. Hair follicle morphogenesis results in the appearance of fur in animals. Increasing evidence suggests that bone morphogenetic proteins (BMP) play an important role in the induction and progression to various stages of the hair follicle cycle. In this study, BMP2 mRNA expression level was investigated from different phases using real-time PCR (qRT-PCR) and localized with in situ hybridization in adult Inner Mongolia Cashmere goat. qRT-PCR revealed that the expression of BMP2 in early anagen was 27.8 times greater than expression during late anagen phase. The in situ hybridization detected BMP2 expressed at days 75 and 115 in both primary and secondary hair follicles during the embryonic period. Further, BMP 2 was expressed in primary hair follicles in late telogen and in secondary hair follicles in early anagen in adult goat. However, it was found to be absent in bo...

SNP Discovery from Transcriptome of Cashmere Goat Skin

The goat Capra hircus is one of several economically important livestock in China. Advances in molecular genetics have led to the identification of several single nucleotide variation markers associated with genes affecting economic traits. Validation of single nucleotide variations in a whole-transcriptome sequencing is critical for understanding the information of molecular genetics. In this paper, we aim to develop a large amount of convinced single nucleotide polymorphisms (SNPs) for Cashmere goat through transcriptome sequencing. In this study, the transcriptomes of Cashmere goat skin at four stages were measured using RNA-sequencing and 90% to 92% unique-mapped-reads were obtained from total-mapped-reads. A total of 56,231 putative SNPs distributed among 10,057 genes were identified. The average minor allele frequency of total SNPs was 18%. GO and KEGG pathway analysis were conducted to analyze the genes containing SNPs. Our follow up biological validation revealed that 64% of SNPs were true SNPs. Our results show that RNA-sequencing is a fast and efficient method for identification of a large number of SNPs. This work provides significant genetic resources for further research on Cashmere goats, especially for the high density linkage map construction and genome-wide association studies.

Enumeration and strain characterization of fecal Escherichia coli associated with feeding triticale dried distillers grain with solubles in beef cattle diets.

Triticale dried distillers grain with solubles (TDDGS), a major by-product of the bioethanol industry, has potential for utilization in animal feed. This study investigated the changes in generic fecal Escherichia coli strains associated with inclusion of TDDGS in cattle diets. Within this study, a longitudinal experiment (112 days) examined the effect of step-up increasing TDDGS inclusion from control to a final diet containing 30% TDDGS among cattle (n = 4), and a short-term experiment (28 days) compared animals (n = 16) fed control, 20%, 25%, or 30% TDDGS diets. We found that incorporation of either 20%, 25%, or 30% TDDGS did not have any effect on the amount of total E. coli shedding over either the longitudinal (p = 0.06) or the short-term (p = 0. 87) study. In both the experiments, 67% of the total E. coli isolates were found to be resistant to one or more of the 17 antimicrobials tested. Among the resistant isolates, cephalothin was the most prevalent resistance (44% isolates). Over the duration of the study, tet(C) was a commonly detected resistance gene in tetracycline-resistant E. coli. Significant diversity was observed among isolates with 33 and 31 pulsed-field gel electrophoresis patterns clustering into 11 and 10 restriction endonuclease digestion pattern clusters for the longitudinal and short-term studies, respectively. Neither the duration of feeding nor increasing the proportion of TDDGS within the diet affected the diversity of E. coli resistance phenotypes or the clonal relatedness of the observed strains. Individual animals retained similar or closely related strains. Based on this study, inclusion of TDDGS as a protein and fiber source in cattle diets is not associated with increased maintenance, shedding, or proliferation of resistant strains of generic E. coli, which is an important reservoir of antimicrobial resistance among cattle.

Expression of Vimentin in hair follicle growth cycle of inner Mongolian Cashmere goats

BackgroundThe growth of Inner Mongolian Cashmere goat skin hair follicle exhibits a periodic growth pattern. The hair growth cycle is distinguished as telogen, anagen, and catagen stages. The role of vimentin in the growth process of hair follicles is evident. To elucidate the mechanism underlying the vimentin activity in the growth cycle of hair follicles, transcriptome sequencing and liquid chromatography-tandem mass spectrometry were used to obtain the nucleic acid and amino acid sequences of VIIM gene and vimentin. The amino acid and nucleic acid sequences were analyzed by comparison. Real-time quantitative PCR, Western blot, and immunohistochemistry analyzed the expression level and sites of vimentin in the three growth stages of the Inner Mongolia Cashmere goat skin samples.ResultsVIM gene cDNA, obtained by transcriptome sequencing, was aligned against that of the Capra hircus VIM gene. The amino acid sequence of vimentin revealed a high similarity rate across other species. The expressions of both VIM gene and vimentin were highest during the growth period and lowest in the rest period. Furthermore, vimentin was primarily expressed in the outer root sheath of the hair follicle as assessed by staining.ConclusionsThe sequences of the gene and protein are similar to that of other species and identical to Capra hircus. However, the expression of VIM and vimentin was proportional to that of the growth of hair follicles. And vimentin expressed only in the outer root sheath of hair follicles. Thus, vimentin was speculated to participate in the regulation of the hair follicle growth cycle by affecting the outer root sheath.

Expression of selenium-independent glutathione peroxidase 5 (GPx5) in the epididymis of Small Tail Han sheep

Objective Selenium-independent glutathione peroxidase (GPx5) is specifically expressed in the mammalian epididymis and plays an important role in protecting sperm from reactive oxygen species and lipid peroxidation damage. This study investigates GPx5 expression in the epididymis of Small Tail Han sheep. Methods GPx5 expression was studied in three age groups: lamb (2 to 3 months), young (8 to 10 months), and adult (18 to 24 months). The epididymis of each age group divided into caput, corpus and cauda, respectively. Analysis the expression quantity of GPx5 in epididymis and testis by real-time fluorescent quantitative polymerase chain reaction and Western blot. Finally, GPx5 protein locating in the epididymis by immunohistochemical. Results The results demonstrate that in the lamb group, the GPx5 mRNA, but not protein, can be detected. GPx5 mRNA and expressed protein were detected in both the young and adult groups. Moreover, both the mRNA and protein levels of GPx5 were significantly higher in the young group than in other two groups. When the different segments of epididymis were investigated, GPx5 mRNA was expressed in each segment of epididymis regardless of age. Additionally, the mRNA level in the caput was significantly higher than that in corpus and cauda within same age group. The GPx5 protein was in the epithelial cells’ cytoplasm. However, GPx5 mRNA and protein were not detected in the testis. Conclusion These results suggest that GPx5 is mainly expressed in the epididymis of Small Tail Han sheep, and that the expression level of GPx5 is associated with age. Additionally, GPx5 was primarily expressed in the epithelial cells of the caput. Taken together, these studies indicate that GPx5 is expressed in the epididymis in all age grades.

High-Throughput Sequencing of Hair Follicle Development-Related Micrornas in Cashmere Goat at Various Fetal Periods

Inner Mongolia cashmere goat marks a precious gerplasm genetic resource due to its excellent cashmere traits. Therefore, it is of crucial importance to investigate the cashmere development mechanism of cashmere goat and to search for the important cashmere growth-related candidate genes. Fetal skin samples at 10 different periods of cashmere goat were collected in this research. Moreover, high-throughput sequencing was conducted on RNA samples from side skin of cashmere goat fetuses collected at three critical periods of skin hair follicle initiation, growth and development (namely, 45, 55 and 65 days) after balanced mix in line with the previous research results. Meanwhile, 3 samples at corresponding periods were used as the biological duplications. Data regarding microRNA and mRNA expression in skin and hair follicles of cashmere goats at various fetal periods were obtained using the high-throughput sequencing method. The results indicated that microRNAs in the oar-let-7 and oar-miR-200 families in 55 days and 66 days of pregnancy samples had been notably up-regulated relative to those in 45 days of pregnancy samples. This revealed that they might be the critical microRNAs in hair follicle development.

Expression of fox-related genes in the skin follicles of Inner Mongolia cashmere goat

Objective This study investigated the expression of genes in cashmere goats at different periods of their fetal development. Methods Bioinformatics analysis was used to evaluate data obtained by transcriptome sequencing of fetus skin samples collected from Inner Mongolia cashmere goats on days 45, 55, and 65 of fetal age. Results We found that FoxN1, FoxE1, and FoxI3 genes of the Fox gene family were probably involved in the growth and development of the follicle and the formation of hair, which is consistent with previous findings. Real-time quantitative polymerase chain reaction detecting system and Western blot analysis were employed to study the relative differentially expressed genes FoxN1, FoxE1, and FoxI3 in the body skin of cashmere goat fetuses and adult individuals. Conclusion This study provided new fundamental information for further investigation of the genes related to follicle development and exploration of their roles in hair follicle initiation, growth, and development.