Characterization of expressed sequence tags from a gallus gallus pineal gland cDNA library

The pineal gland is the circadian oscillator in the chicken, regulating diverse functions ranging from egg laying to feeding. Here, we describe the isolation and characterization of expressed sequence tags (ESTs) isolated from a chicken pineal gland cDNA library. A total of 192 unique sequences were analysed and submitted to GenBank; 6% of the ESTs matched neither GenBank cDNA sequences nor the newly assembled chicken genomic DNA sequence, three ESTs aligned with sequences designated to be on the Z_random, while one matched a W chromosome sequence and could be useful in cataloguing functionally important genes on this sex chromosome. Additionally, single nucleotide polymorphisms (SNPs) were identified and validated in 10 ESTs that showed 98% or higher sequence similarity to known chicken genes. Here, we have described resources that may be useful in comparative and functional genomic analysis of genes expressed in an important organ, the pineal gland, in a model and agriculturally important organism.     

Differential regulation of dihydroceramide desaturase by palmitate versus monounsaturated fatty acids: implications for insulin resistance

Much data implicate saturated fatty acids in deleterious processes associated with obesity, diabetes, and the metabolic syndrome. Many of these changes may be due to aberrant generation of bioactive lipids when saturated fatty acid availability to tissues is increased. On the other hand, studies are emerging that implicate the monounsaturated fatty acid oleate in protection from saturated fat mediated toxicity; however, the mechanisms are not well understood. Our data demonstrate a novel role for palmitate in increasing mRNA encoding DES1, which is the enzyme responsible for generating ceramide from its precursor dihydroceramide and thus controls synthesis of the bioactive lipid ceramide. Moreover, co-treatment with oleate prevented the increase in ceramide, and this occurred through attenuation of the increase in message and activity of DES1. Knockdown of DES1 also protected from palmitate-induced insulin resistance, and overexpression of this enzyme ameliorated the protective effect of oleate. Together, these findings provide insight into the mechanisms of oleate-mediated protection against metabolic disease and provide novel evidence for fatty acid-mediated regulation of a key enzyme of ceramide biosynthesis.     

A copper‐specific microbial fuel cell biosensor based on riboflavin biosynthesis of engineered Escherichia coli

Copper pollution poses a serious threat to the aquatic environment; however, in situ analytical methods for copper monitoring are still scarce. In the current study, Escherichia coli Rosetta was genetically modified to express OprF and ribB with promoter P_t7 and P_cusC, respectively, which could synthesize porin and senses Cu²⁺ to produce riboflavin. The cell membrane permeability of this engineered strain was increased and its riboflavin production (1.45–3.56 μM) was positively correlated to Cu²⁺ (0–0.5 mM). The biosynthetic strain was then employed in microbial fuel cell (MFC) based biosensor. Under optimal operating parameters of pH 7.1 and 37°C, the maximum voltage (248, 295, 333, 352, and 407 mV) of the constructed MFC biosensor showed a linear correlation with Cu²⁺ concentration (0.1, 0.2, 0.3, 0.4, 0.5 mM, respectively; R² = 0.977). The continuous mode testing demonstrated that the MFC biosensor specifically senses Cu²⁺ with calculated detection limit of 28 μM, which conforms to the common Cu²⁺ safety standard (32 μM). The results obtained with the developed biosensor system were consistent with the existing analytical methods such as colorimetry, flame atomic absorption spectrometry, and inductively coupled plasma optical emission spectrometry. In conclusion, this MFC‐based biosensor overcomes the signal conversion and transmission problems of conventional approaches, providing a fast and economic analytical alternative for in situ monitoring of Cu²⁺ in water.     

SphK1 mediates hepatic inflammation in a mouse model of NASH induced by high saturated fat feeding and initiates proinflammatory signaling in hepatocytes

Steatohepatitis occurs in up to 20% of patients with fatty liver disease and leads to its primary disease outcomes, including fibrosis, cirrhosis, and increased risk of hepatocellular carcinoma. Mechanisms that mediate this inflammation are of major interest. We previously showed that overload of saturated fatty acids, such as that which occurs with metabolic syndrome, induced sphingosine kinase 1 (SphK1), an enzyme that generates sphingosine-1-phosphate (S1P). While data suggest beneficial roles for S1P in some contexts, we hypothesized that it may promote hepatic inflammation in the context of obesity. Consistent with this, we observed 2-fold elevation of this enzyme in livers from humans with nonalcoholic fatty liver disease and also in mice with high saturated fat feeding, which recapitulated the human disease. Mice exhibited activation of NFκB, elevated cytokine production, and immune cell infiltration. Importantly, SphK1-null mice were protected from these outcomes. Studies in cultured cells demonstrated saturated fatty acid induction of SphK1 message, protein, and activity, and also a requirement of the enzyme for NFκB signaling and increased mRNA encoding TNFα and MCP1. Moreover, saturated fat-induced NFκB signaling and elevation of TNFα and MCP1 mRNA in HepG2 cells was blocked by targeted knockdown of S1P receptor 1, supporting a role for this lipid signaling pathway in inflammation in nonalcoholic fatty liver disease.     

微卫星技术分析地方鸡种群体的遗传结构

利用微卫星技术和14对引物MCW150、MCW134、MCW145、MCW104、ADL210、MCW88、ADL237、ADL201、ADL289、MCW5、MCW217、MCW29、ADL176、MCW4对7个地方鸡种和1个引进鸡种的群体遗传结构特征进行了分析。根据池DNA所获得的品种间相似指数计算遗传距离。结果表明:AA鸡与其它品种鸡遗传距离最远,白耳鸡次之,其余品种间较近。这些鸡种的遗传结构特征与其地理分布有关,说明微卫星技术作为畜禽遗传结构分析的辅助手段是可行的,也是高效的 。     

Bulge oligonucleotide as an inhibitory agent of bacterial topoisomerase I

Bacterial topoisomerase I (Btopo I) was defined as potential target for discovery of new antibacterial compounds. Various oligonucleotides containing bulge structure were designed and synthesised as inhibitors to Btopo I in this investigation. The results of this study demonstrated that the designed oligonucleotides display high inhibitory efficiency on the activity of Btopo I and the inhibitory effect could be modulated by the amount of bulge DNA bases. The most efficient one among them showed an IC₅₀ value of 63.1?nM in its inhibition on the activity of Btopo I. In addition, our studies confirmed that the designed oligonucleotide would induce irreversible damages to Btopo I and without any effects occur to eukaryotic topoisomerase I. It is our hope that the results provided in these studies could provide a novel way to inhibit Btopo I.     

DNA sequence and haplotype variation in two candidate genes for dilated cardiomyopathy in the turkey Meleagris gallopavo

Determining variation in genes is fundamental to understanding their function in the disease state. Cardiac troponin T (cTnT) and phospholamban (PLN) genes have been implicated in dilated cardiomyopathy (DCM) in human and model species. To investigate the role of these 2 candidate genes in DCM in the turkey Meleagris gallopavo, understanding sequence variants and map position distribution is necessary. To this end, a total of 1854 and 1771 bp of cTnT and PLN gene sequences, respectively, were scanned for single nucleotide polymorphisms (SNPs) in a randomly bred population. A total of 15 SNPs was identified in the cTnT and PLN genomic sequences. Nine haplotypes, 5 in cTnT and 4 in PLN, were identified. Observed heterozygosities (0.02-0.39) in the turkey population were low for both genes. Within each gene, 1 SNP corresponding to a restriction enzyme site was identified and used to develop a PCR-restriction fragment length polymorphism (RFLP) genotyping assay. The PLN gene was genetically mapped to turkey chromosome 2, equivalent to Gallus gallus chromosome 3, and cTnT mapped to a turkey microchromosome. Although limited because of the relatively small sample size of 55 birds, the data from this SNP analysis of PLN and cTnT provide a foundation from which to evaluate the function of cTnT and PLN in the turkey. Information about the distribution of the SNPs and haplotypes will facilitate future association and linkage studies.     

Generation of Cas9 transgenic zebrafish and their application in establishing an ERV-deficient animal model

Objectives

To investigate the effect of endogenous Cas9 on genome editing efficiency in transgenic zebrafish.

Results

Here we have constructed a transgenic zebrafish strain that can be screened by pigment deficiency. Compared with the traditional CRISPR injection method, the transgenic zebrafish can improve the efficiency of genome editing significantly. At the same time, we first observed that the phenotype of vertebral malformation in early embryonic development of zebrafish after ZFERV knockout.

Conclusions

The transgenic zebrafish with expressed Cas9, is more efficient in genome editing. And the results of ZFERV knockout indicated that ERV may affect the vertebral development by Notch1/Delta D signal pathway.