Xianbo Jia

A transcriptome atlas of rabbit revealed by PacBio single-molecule long-read sequencing.

It is widely acknowledged that transcriptional diversity largely contributes to biological regulation in eukaryotes. Since the advent of second-generation sequencing technologies, a large number of RNA sequencing studies have considerably improved our understanding of transcriptome complexity. However, it still remains a huge challenge for obtaining full-length transcripts because of difficulties in the short read-based assembly. In the present study we employ PacBio single-molecule long-read sequencing technology for whole-transcriptome profiling in rabbit (Oryctolagus cuniculus). We totally obtain 36,186 high-confidence transcripts from 14,474 genic loci, among which more than 23% of genic loci and 66% of isoforms have not been annotated yet within the current reference genome. Furthermore, about 17% of transcripts are computationally revealed to be non-coding RNAs. Up to 24,797 alternative splicing (AS) and 11,184 alternative polyadenylation (APA) events are detected within this de novo constructed transcriptome, respectively. The results provide a comprehensive set of reference transcripts and hence contribute to the improved annotation of rabbit genome.

Association of a synonymous mutation of the PGAM2 gene and growth traits in rabbits

Phosphoglycerate mutase (PGAM2) catalyzes the conversion of 3-phosphoglycerate into 2-phosphoglycerate and releases energy during glycolysis in muscle tissues. PGAM2 has been considered as a candidate gene to influence growth, development, and carcass traits in livestock. The aim of this study was to investigate the association between polymorphisms of PGAM2 and growth traits in rabbits. Three single nucleotide polymorphisms (SNPs) were identified by direct sequencing in 20 random individuals from three breeds, including c.-10C>T, c.195C>T, and c.414+17C>T. The c.195C>T was genotyped by PCR-RFLP in a total of 222 rabbits of three breeds (Tianfu black, 53 animals; Ira, 91 animals; Champagne, 78 animals). The average allele frequency among the breeds was 0.52 for allele T and 0.48 for C. The heterozygosity and effective number of alleles were 0.4992 and 1.996, respectively. The association results revealed the CT genotype of c.195C>T was associated significantly (P < 0.05) with greater body weight at 84 days of age (BW84) and with average daily weight gain (ADG). However, association of the genotypes with other production traits was not observed. The results of this study suggested PGAM2 is one of the candidate genes affecting BW84 and ADG in the rabbit.

A novel single nucleotide polymorphism of the POU1F1 gene associated with meat quality traits in rabbits

Abstract The purpose of this research was to investigate the effect of the POU1F1 gene on meat quality traits in the Hyla, Champagne, and Tianfu Black rabbit breeds. We detected one single nucleotide polymorphism and the SNP was located at 536 bp in intron 5 of this gene. Chi-square tests showed that the genotypic frequencies in the three rabbit populations were not in Hardy-Weinberg equilibrium. The PIC values indicated that the three populations had intermediate levels of genetic diversity. Rabbits with the CC genotype had a significantly greater pH0h than those with the CT genotype in the biceps femoris muscle. The least squares means for cooking loss in CT and CC rabbits were significantly higher than those for TT rabbits. Rabbits with the CC genotype had a significantly higher intramuscular fat content in the longissimus dorsi and biceps femoris muscles than those with genotype TT and CT. Thus, the results here indicate that this POU1F1 SNP may be of potential use in marker assisted selection for meat quality traits in rabbits.

bioOTU: An Improved Method for Simultaneous Taxonomic Assignments and Operational Taxonomic Units Clustering of 16s rRNA Gene Sequences

Clustering of 16s rRNA amplicon sequences into operational taxonomic units (OTUs) is the most common bioinformatics pipeline for investigating microbial community by high-throughput sequencing technologies. However, the existing algorithms of OTUs clustering still remain to be improved at reliability. Here we propose an improved method (bioOTU) that first assigns taxonomy to unique tags at genus level for separating the error-free sequences of known species in reference database from artifacts, and then cluster them into OTUs by different strategies. The remaining tags, which fail to be clustered in the previous step, are further subjected to independent OTUs clustering by the optimized algorithm of heuristic clustering. The performance tests on both mock and real communities revealed that bioOTU is powerful for recovering the underlying profiles at both microbial composition and abundance, and it also produces comparable or less number of OTUs in comparison with the prevailing tools of Mothur and UPARSE. The bioOTU is implemented in C and Python languages with source codes freely available on the GitHub repository.

miR-221 modulates skeletal muscle satellite cells proliferation and differentiation

MicroRNAs (miRNAs) are a class of small non-coding RNA molecules, which play important roles in animals by targeting mRNA transcripts for translational repression. Many recent studies have shown that miRNAs are involved in the control of muscle development. In this study, the expression levels of miR-221 in different tissues and during rabbit skeletal muscle satellite cells (SMSCs) differentiation were detected. Gene ontology term enrichment was used to predict the potential biological roles of miR-221. A synthetic miR-221 mimic and a miR-221 inhibitor were used to investigate the functions of miR-221 during SMSCs proliferation and differentiation to further verify the functions of miR-221 in muscle development. In this report, we compared the expression levels of miR-221 in different tissues. The expression levels of miR-221 were upregulated after the induction of differentiation, and then were gradually downregulated during SMSCs differentiation. Overexpression of miR-221 promoted SMSCs proliferation, whereas inhibiting expression restrained proliferation in the EdU and CCK-8 assays. In addition, overexpression of miR-221 led to a decline in the expression levels of the differentiation marker genes MyoG and MHC. miR-221 overexpression suppressed SMSCs myotube formation. On the contrary, inhibition of miR-221 promoted myotube formation. Our data showed that miR-221 increased SMSCs proliferation and decreased differentiation.

Genetic effect of Myf5 gene in rabbit meat quality traits

The objective of this study was to investigate the effect of the polymorphism in the Myf5 gene on meat quality traits in the Ira and Tianfu Black rabbit breeds using polymerase chain reaction and DNA sequencing. A total of six SNPs and four haplotypes were found in Ira rabbits and only two SNPs were found in Tianfu Black rabbits. The two rabbit breeds had intermediate levels of genetic diversity according to their polymorphic information content values. The SNP association analysis in Ira indicated that SNP1-6 had a significant association with redness, yellowness and intramuscular fat values in the biceps femoris muscle, and also a significantly effect on redness in the longissimus dorsi muscle. The haplotype association analysis indicated that some haplotypes could be selected to get higher or lower meat redness, yellowness and intramuscular fat content in longissimus dorsi in Ira rabbits. Several SNPs and haplotypes of Myf5 identified here could be considered as molecular markers to improve the meat quality of Ira and Tianfu Black rabbits.

Calpastatin gene polymorphism is associated with rabbit meat quality traits

ABSTRACT The objective of this study was to investigate single nucleotide polymorphisms in the calpastatin (CAST) gene and to test their association with meat quality traits in Hyla, Champagne, and Tianfu Black rabbit breeds. We detected one single nucleotide polymorphism (g.16441502 C  > T) located at 67 bp in intron 3 of the CAST gene in Chromosome 11. The three rabbit populations had intermediate levels of genetic diversity in the CAST gene. The statistical analysis indicated that rabbits with the TT genotype had a significantly greater yellowness at 0 and 24 h postmortem than those with the CC genotype (P < .05) in the longissimus dorsi muscle. Rabbits with CC genotype had higher intramuscular fat content than those with CT and CC genotypes in both longissimus dorsi and biceps femoris muscles in the three breeds (P < .05). Our results indicated that the CAST gene may be used as a possible candidate for marker-assisted selection in rabbit meat breeding programmes.

Investigation of JAKs/STAT-3 in lipopolysaccharide-induced intestinal epithelial cells

Janus‐activated kinase (JAKs)‐signal transducer and activator of transcription 3 (STAT‐3) signalling play critical roles in immunoregulation and immunopathology, which involve inflammatory responses and enteritis. JAK phosphorylates STAT‐3 in response to stimulation by cytokines or growth factors, and then activates or represses the gene expression. STAT‐3 is activated persistently in cancer cells and contributes to the malignant progression of various types of cancer and inflammation. To elucidate the different roles of JAKs in the activation of STAT‐3, the lipopolysaccharide‐induced primary intestinal epithelial cell (IEC) acute inflammatory model was established. Small interference RNAs (siRNAs) were then employed to attenuate the expression levels of JAKs. Real‐time quantitative reverse transcription–polymerase chain reaction (PCR) (qRT–PCR) revealed that JAK mRNA levels were reduced efficiently by JAK‐specific siRNAs. Under the IEC inflammatory model transfected with si‐JAK, which equates to effective silencing, qRT–PCR and Western blot assays, suggested that knockdowns of JAK attenuated the JAK‐induced down‐regulation of STAT‐3 at the mRNA or protein levels. In particular, JAK1 played a key role, which was consistent with the RNA‐Seq results. Subsequently, the expression levels of proinflammatory cytokines interleukin (IL)‐1β and tumour necrosis factor (TNF)‐α were down‐regulated in the IEC inflammatory model transfected with si‐JAK1. JAK1 appears as a direct activator for STAT‐3, whereas treatments targeting JAK1 repressed STAT‐3 sufficiently pathways in the IEC inflammatory model. Therefore, the control of JAK1 using siRNAs has the potential to be an effective strategy against enteritis.

The novel SNPs of leptin gene and their associations with growth traits in Chinese Nanjiang Yellow goat.

The leptin (LEP) gene encodes a protein that greatly affects the regulation of body weight, energy balance, and food intake in mammals. The objective of the present work was to identify genetic variants of the caprine LEP gene in 411 individuals from five Chinese goat breeds. Six novel single nucleotide polymorphisms (SNPs) (g.117T > C, g.1642G > A, g.2883G > A, g.3053T > C, g.3190G > A, and g.3314T > C) were detected using DNA sequencing. A chi-squared (χ(2)) test showed that all of the LEP SNPs were in Hardy-Weinberg equilibrium in the studied population (P > 0.05). Six common haplotypes were identified in the five goat populations, with frequencies ranging from 0.083 to 0.244. The r(2) linkage disequilibrium plot of the LEP SNPs indicated linkage disequilibrium only in the cultured breeds (NJ and JY). Statistical analysis revealed that all of the six SNPs of the LEP gene were associated with growth traits. The individuals with the GG genotype at g.1642G>A and g.3190G > A loci showed higher birth weight (2.38 ± 0.03, 2.43 ± 0.05) and weight at 2 months of age (10.59 ± 0.16, 10.71 ± 0.26) than the A-bearing genotypes (AA or GA, P < 0.05). Our findings indicate that polymorphisms of the caprine LEP gene might be important genetic factors influencing growth traits, and these genetic markers may be useful for future marker-assisted selection programs in goat breeding and production.

Effects of Variants in Proopiomelanocortin and Neuropeptide Y Genes on Growth, Carcass, and Meat Quality Traits in Rabbits

Appetite-related neuropeptides proopiomelanocortin (POMC) and Neuropeptide Y (NPY) are essential for regulating feeding behavior and energy homeostasis. The objective of this study was to evaluate the effects of variants in POMC and NPY genes on growth, carcass and meat quality traits in rabbits. A total of six SNPs were identified for POMC (n = 2) and NPY (n = 4) genes by direct sequencing. Three SNPs were subsequently genotyped by using MassArray system (Sequenom iPLEXassay) in 235 individuals, which belong to three meat rabbit breeds, including 93 Ira rabbits; 81 Champagne rabbits and 61 Tianfu black rabbits. The SNP c.112-12G>T was in intron-exon boundaries (intron 1) of POMC gene, and the association analysis showed that individuals with TT genotype had a greater 84 d body weight (BW84), eviscerated weight and semi-eviscerated weight than those with GT genotype (p<0.05); the TT individuals were also higher than those GG in the ripe meat ratio (RMR) (p<0.05). The g.1778G>C SNP, which was in complete linkage with other three SNPs (g.1491G>A, g.1525G>T and g.1530C>T) in intron 1 of NPY gene, was significantly correlated with eviscerated slaughter percentage and semi-eviscerated slaughter percentage in rabbits, and the individuals with CC genotype had a better performance than CG genotype (p<0.05). These findings would provide primary clues for the biological roles of POMC and NPY underlying the rabbit growth-related traits.