传染性法氏囊病病毒在次代鸡胚成纤维细胞上的增殖

研究了用次代鸡胚成纤维细胞（SCEF）增殖传染性法氏囊病病毒（IBDV）的可能性。在研究了原代鸡胚成纤维细胞（PCEF）与SCEF的生长特性的基础上,就各种培养方式采用PCEF与SCEF增殖IBDV进行了比较。结果表明,可用SCEF代替PCEF进行IBDV的增殖培养。     

Efficient assembly of a large fragment of monkeypox virus genome as a qPCR template using dual-selection based transformation-associated recombination

Transformation-associated recombination (TAR) has been widely used to assemble large DNA constructs. One of the significant obstacles hindering assembly efficiency is the presence of error-prone DNA repair pathways in yeast, which results in vector backbone recircularization or illegitimate recombination products. To increase TAR assembly efficiency, we prepared a dual-selective TAR vector, pGFCS, by adding a P_ADH1-URA3 cassette to a previously described yeast-bacteria shuttle vector, pGF, harboring a P_HIS3-HIS3 cassette as a positive selection marker. This new cassette works as a negative selection marker to ensure that yeast harboring a recircularized vector cannot propagate in the presence of 5-fluoroorotic acid. To prevent pGFCS bearing ura3 from recombining with endogenous ura3-52 in the yeast genome, a highly transformable Saccharomyces cerevisiae strain, VL6-48B, was prepared by chromosomal substitution of ura3-52 with a transgene conferring resistance to blasticidin. A 55-kb genomic fragment of monkeypox virus encompassing primary detection targets for quantitative PCR was assembled by TAR using pGFCS in VL6-48B. The pGFCS-mediated TAR assembly showed a zero rate of vector recircularization and an average correct assembly yield of 79% indicating that the dual-selection strategy provides an efficient approach to optimizing TAR assembly.     

Performance of distance-based DNA barcoding in the molecular identification of Primates

For comparative primatology proper recognition of basal taxa (i.e. species) is indispensable, and in this the choice of a suitable gene with high phylogenetic resolution is crucial. For the goals of species identification in animals, the cytochrome c oxidase subunit 1 (cox1) has been introduced as standard marker. Making use of the difference in intra- and interspecific genetic variation – the DNA barcoding gap – cox1 can be used as a fast and accurate marker for the identification of animal species. For the Order Primates we compare the performance of cox1 (166 sequences; 50 nominal species) in species-identification with that of two other mitochondrial markers, 16S ribosomal RNA (412 sequences, 92 species) and cytochrome b (cob: 547 sequences, 72 species). A wide gap exist between intra- and interspecific divergences for both cox1 and cob genes whereas this gap is less apparent for 16S, indicating that rRNA genes are less suitable for species delimitation in DNA barcoding. For those species where multiple sequences are available there are significant differences in the intraspecific genetic distances between different mitochondrial markers, without, however, showing a consistent pattern. We conclude that cox1 allows accurate differentiation of species and as such DNA barcoding may have an important role to play in comparative primatology.     

Viroporins

Viroporins are a group of proteins that participate in several viral functions, including the promotion of release of viral particles from cells. These proteins also affect cellular functions, including the cell vesicle system, glycoprotein trafficking and membrane permeability. Viroporins are not essential for the replication of viruses, but their presence enhances virus growth. Comprising some 60-120 amino acids, viroporins have a hydrophobic transmembrane domain that interacts with and expands the lipid bilayer. Some viroporins also contain other motifs, such as basic amino acid residues or a domain rich in aromatic amino acids that confers on the protein the ability to interact with the interfacial lipid bilayer. Viroporin oligomerization gives rise to hydrophilic pores at the membranes of virus-infected cells. As the list of known viroporins steadily grows, recent research efforts focus on deciphering the actions of the viroporins poliovirus 2B, alphavirus 6K, HIV-1 Vpu and influenza virus M2. All these proteins can enhance the passage of ions and small molecules through membranes depending on their concentration gradient. Future work will lengthen the list of viroporins and will provide a deeper understanding of their mechanisms of action.     

Alisol A attenuates high‐fat‐diet‐induced obesity and metabolic disorders via the AMPK/ACC/SREBP‐1c pathway

Obesity and its associated metabolic disorders such as diabetes, hepatic steatosis and chronic heart diseases are affecting billions of individuals. However there is no satisfactory drug to treat such diseases. In this study, we found that alisol A, a major active triterpene isolated from the Chinese traditional medicine Rhizoma Alismatis, could significantly attenuate high‐fat‐diet‐induced obesity. Our biochemical detection demonstrated that alisol A remarkably decreased lipid levels, alleviated glucose metabolism disorders and insulin resistance in high‐fat‐diet‐induced obese mice. We also found that alisol A reduced hepatic steatosis and improved liver function in the obese mice model.In addition, protein expression investigation revealed that alisol A had an active effect on AMPK/ACC/SREBP‐1c pathway. As suggested by the molecular docking study, such bioactivity of alisol A may result from its selective binding to the catalytic region of AMPK.Therefore, we believe that Alisol A could serve as a promising agent for treatment of obesity and its related metabolic diseases.     

CRISPR/Cas系统在抗病毒研究中的应用

近年来,CRISPR/Cas系统因其效率高、靶向性强、易操作等优势,已被广泛应用于多种病毒研究中。本文首先简单介绍了CRISPR/Cas系统的分类,并比较了Cas9和Cas12a与Cas13a的特点;其次重点介绍了CRISPR/Cas9通过靶向破坏病毒基因组,或编辑宿主关于病毒生命周期的关键因子的策略在抗病毒方面的各种应用,CRISPR/Cas13a采用靶向破坏病毒基因组方法在抗病毒中的应用,以及CRISPR/Cas12a和CRISPR/Cas13a在病毒基因检测中的应用。最后讨论了CRISPR/Cas在病毒研究中面临的挑战,并讨论了CRISPR/Cas12a作为抗病毒工具的潜在应用前景。由于CRISPR/Cas系统自身的优势,预计该系统将会给病毒相关的疾病诊断和控制带来革命性的变化。