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We present the finding of a dimeric ACE2 peptide mimetic designed through side chain cross-linking and covalent dimerization. It has a binding affinity of 16 nM for the SARS-CoV-2 spike RBD, and effectively inhibits the SARS-CoV-2 pseudovirus in Huh7-hACE2 cells with an IC

As one of the major approaches in combating the COVID-19 pandemics, the availability of specific and reliable assays for the SARS-CoV-2 viral genome and its proteins is essential to identify the infection in suspected populations, make diagnoses in symptomatic or asymptomatic individuals, and determine clearance of the virus after the infection. For these purposes, use of the quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) for detection of the viral nucleic acid remains the most valuable in terms of its specificity, fast turn-around, high-throughput capacity, and reliability. It is critical to update the sequences of primers and probes to ensure the detection of newly emerged variants. Various assays for increased levels of IgG or IgM antibodies are available for detecting ongoing or past infection, vaccination responses, and persistence and for identifying high titers of neutralizing antibodies in recovered individuals. Viral genome sequencing is increasingly used for tracing infectious sources, monitoring mutations, and subtype classification and is less valuable in diagnosis because of its capacity and high cost. Nanopore target sequencing with portable options is available for a quick process for sequencing data. Emerging CRISPR-Cas-based assays, such as SHERLOCK and AIOD-CRISPR, for viral genome detection may offer options for prompt and point-of-care detection. Moreover, aptamer-based probes may be multifaceted for developing portable and high-throughput assays with fluorescent or chemiluminescent probes for viral proteins. In conclusion, assays are available for viral genome and protein detection, and the selection of specific assays depends on the purposes of prevention, diagnosis and pandemic control, or monitoring of vaccination efficacy., During the COVID-19 pandemics, sensitive and reliable assays for SARS-CoV-2 detection are essential for screening the population, identifying asymptomatic individuals, making diagnoses, monitoring treatment responses, and determining viral clearance. This review summarizes the principles, advantages, disadvantages, and specific applications of currently available assays for detection of the viral nucleotide, genome or proteins, as well as host antibody responses, and provide overall guidelines for selection of optimal assays for specific usage.

Chronic hepatitis B virus (HBV) infection causes hepatitis, liver cirrhosis and hepatocellular carcinoma. Covalently closed circular DNA (cccDNA) is the transcription template for HBV RNAs and not affected by current treatment options. Effective therapeutics with ability to remove cccDNA need to be developed. Previously, we established an HBV persistence mouse model via hydrodynamic injection of a clinical isolate (BPS) and identified IL-21 as a potent inducer of viral clearance. Here, we aimed to explore the anti-HBV effects of IL-21 messenger RNA (mRNA) delivered by lipid nanoparticle (LNP-IL-21) system. First, LNP-IL-21 was prepared and analyzed for its safety, expression, biodistribution and stability in vitro and in vivo. Next, LNP-IL-21 was injected into two HBV persistence mouse models based on BPS and recombinant cccDNA (rcccDNA) respectively. LNP-IL-21 administration successfully cleared HBV serum markers, and more importantly, BPS replicons and rcccDNA in livers, which was associated with activation of viral specific immune responses. Notably, transfer of peripheral blood mononuclear cells from BPS persistence mice stimulated ex vivo with LNP-IL-21 and viral antigen could induce HBV clearance in recipient mice. These findings suggested that both LNP-IL-21-based gene and cellular therapies provided novel therapeutic strategies against chronic HBV infection.

There is a potential risk for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spread through human contact with seafood and the inanimate materials contaminated by the virus. In this study, we examined the stability of the virus in artificial seawater (ASW) and on the surface of selected materials. SARS-CoV-2 (3.75 log

As a heat treatable aluminum alloy to be used in T6 and T8 temper, belongs to Al-Cu-Li system, a novel high-strength aluminum-lithium alloy 2A97 was developed. In order to improve the relationships of strength and ductility and fracture toughness, and to urge the applications in the aeronautical and aerospace industries, the effects of normal heat treatments and thermomechanical heat treatments on the mechanical properties and fracture toughness were investigated by Transmission Electron Microscope(TEM), Scanning Electronic Microscope (SEM), tensile test, and fracture toughness test. The results show that for the alloy aged at 135 °C for 24 h after quenching and 4 percent plastic deformation, its microstructures are strengthened by strain hardening and precipitation hardening, consisting of fine T1phase, θ″/θ′ phase and δ′ phase densely and homogeneously distributed in the matrix. It yields optimum relationship of strength and ductility, fracture toughness, its σ0.2, σband δ5are 454 MPa, 536 MPa, and 11.8%, respectively. It yields 43.5 MPa·m1/2of Kqvalues higher than that of 42.5 MPa·m1/2 in T6 temper. The fracture morphologies of impact tensile samples of fracture toughness test and normal tensile test were observed, indicating the dominance of intergranular failure and subintergranular failure with some dimples and trangranular failure.

Retrogression characteristics of a novel Al-Cu-Li-X alloy of 2A97 were studied by hardness testing, transmission electron microscopy (TEM), and differential scanning calorimetry (DSC). The retrogression treatments of aging at 155°C for 12 h followed by aging at 220 and 240°C were chosen by determining the peak temperature of δ′ precipitation at 230°C by DSC. The retrogression treatment at a lower temperature of 220°C causes the precipitation and coarsening of δ′ and θ′ phases in the matrix, resulting in an increase in hardness. Retrogression at a higher temperature of 240°C causes the dissolution and coarsening of δ′ and θ′ precipitates in the matrix and on the grain boundaries, resulting in a decrease in hardness. Microstructural changes upon retrogression including the appearance of equilibrium precipitates such as T1, T2, δ′, and θ are confirmed by the selected area electron diffraction and the bright and dark field image analysis.

In hepatitis B virus (HBV) replication cycle, pregenomic RNA undergoes splicing and the reverse transcribed defective genomes can be packaged and released. Various types of spliced defective HBV genomes have been isolated from the sera and liver tissues of viral hepatitis B patients. To explore the functions of a 2.2 kb double spliced HBV variant, a 3.2 kb full-length HBV isolate (#97–34) and its 2.2 kb double-spliced HBV variant (#AP–12) from the tumor tissue of a patient with hepatocellular carcinoma (HCC) were amplified and cloned. Sequencing results showed that #AP12 had deletions in pre-S2, part of pre-S1, S genes, part of the spacer, and part of the reverse transcriptase gene, while the X gene was intact. When this defective double-spliced genome and its full-length counterpart genome were co-transfected into HepG2 cells, the former was shown to enhance the replication of the latter, both by real-time PCR and Southern blotting. When a replication incompetent clone 97–34G1881A was used to co-transfect with #AP12, #AP12 DNA was increased, indicating that replication of the wild-type virus was not the only factor involved in this observation. However, the replication enhancing competency of #AP12 was shown to require an intact HBV X expression cassette. The double-spliced defective variant might contribute to persistent HBV replication in a subpopulation of HCC patients. J. Med. Virol. 81:230–237, 2009. © 2008 Wiley-Liss, Inc.

Recently, a new super-high-strength Al-Zn-Mg-Cu alloy with Zr bearing was developed in BIAM. In this paper, the effect of microalloying element Zr on the microstructure and mechanical properties of the alloy was investigated. It was found that the influence of Zr on the microstructure and age-hardening behaviors was remarkable. The strength of the alloy increases with increasing the content of zirconium, and the peak value of the elongation appeared when the content of Zr was 0.06%(mass fraction) and then decreases. The fracture behavior was also studied. The result shows that the fracture mechanism of the alloy was converted from intergranular brittle to transgranular ductile model with increasing the content of zirconium. Finally, the optimum contents of Zr in the alloy were proposed as 0.10%~0.14%.

A new high strength 2A97 Al-Cu-Li-X alloy was subjected to triple-aging of retrogression and re-aging treatments (RRA). Transmission electron microscopy (TEM), differential scanning calorimetry (DSC), and tensile tests were used to investigate the effects of RRA treatment on the microstructures and properties. DSC test reveals the reversion temperature range of the strengthening δ' (Al3Li) phase. The results show that the microstructure consists of δ' (Al3Li) phase, T1 (Al2CuLi) phase and θ″/θ'(Al2Cu) phase for 2A97 alloy treated by a triple-aging of a retrogression and re-aging treatment in the following order: (1) at 165 °C× 30 min, (2) at 220 °C or 240 ° × 15 min, (3) at 165 °C× 24 h. The plastic deformation, incorporated into the treatment after secondary high temperature aging, promotes the T1 precipitation during final re-aging. The tensile properties of the alloy treated by the retrogression and re-aging treatment reach the peak level of alloy single-aged at 165 °C in T6 temper.

Spike protein is one of the major structural proteins of severe acute respiratory syndrome-coronavirus. It is essential for the interaction of the virons with host cell receptors and subsequent fusion of the viral envelop with host cell membrane to allow infection. Some spike proteins of coronavirus, such as MHV, HCoV-OC43, AIBV and BcoV, are proteolytically cleaved into two subunits, S1 and S2. In contrast, TGV, FIPV and HCoV-229E are not. Many studies have shown that the cleavage of spike protein seriously affects its function. In order to investigate the maturation and proteolytic processing of the S protein of SARS CoV, we generated S1 and S2 subunit specific antibodies (Abs) as well as N, E and 3CL protein-specific Abs. Our results showed that the antibodies could efficiently and specifically bind to their corresponding proteins from E.coli expressed or lysate of SARS-CoV infected Vero-E6 cells by Western blot analysis. Furthermore, the anti-S1 and S2 Abs were proved to be capable of binding to SARS CoV under electron microscope observation. When S2 Ab was used to perform immune precipitation with lysate of SARS-CoV infected cells, a cleaved S2 fragment was detected with S2-specific mAb by Western blot analysis. The data demonstrated that the cleavage of S protein was observed in the lysate, indicating that proteolytic processing of S protein is present in host cells.

Proteomics was used to identify a protein encoded by ORF 3a in a SARS-associated coronavirus (SARS-CoV). Immuno-blotting revealed that interchain disulfide bonds might be formed between this protein and the spike protein. ELISA indicated that sera from SARS patients have significant positive reactions with synthesized peptides derived from the 3a protein. These results are concordant with that of a spike protein-derived peptide. A tendency exists for co-mutation between the 3a protein and the spike protein of SARS-CoV isolates, suggesting that the function of the 3a protein correlates with the spike protein. Taken together, the 3a protein might be tightly correlated to the spike protein in the SARS-CoV functions. The 3a protein may serve as a new clinical marker or drug target for SARS treatment.

The nucleocapsid (N) protein of severe acute respiratory syndrome-coronavirus (SARS-CoV) is a major virion structural protein. In this study, two epitopes (N1 and N2) of the N protein of SARS-CoV were predicted by bioinformatics analysis. After immunization with two peptides, the peptides-specific antibodies were isolated from the immunized rabbits. The further experiments demonstrated that N1 peptide-induced polyclonal antibodies had a high affinity to bind to E. coli expressed N protein of SARS-CoV. Furthermore, it was confirmed that N1 peptide-specific IgG antibodies were detectable in the sera of severe acute respiratory syndrome (SARS) patients. The results indicated that an epitope of the N protein has been identified and N protein specific Abs were produced by peptide immunization, which will be usefull for the study of SARS-CoV.

Enhancer II (ENII) of hepatitis B virus (HBV) is one of the essential cis-elements for the transcriptional regulation of HBV gene expression. Its function is highly liver-specific, suggesting that liver-enriched transcriptional factors play critical roles in regulating the activity of ENII. In this report, a novel hepatocyte transcription factor, which binds specifically to the B1 region (AACGACCGACCTTGAG) within the major functional unit (B unit) of ENII, has been cloned from a human liver cDNA library by yeast one-hybrid screening, and demonstrated to trans-activate ENII via the B1 region. We named this factor hB1F, for human B1-binding factor. Amino acid analysis revealed this factor structurally belongs to nuclear receptor superfamily. Based on the sequence similarities, hB1F is characterized to be a novel human homolog of the orphan receptor fushi tarazu factor I (FTZ-F1). Using reverse transcription-polymerase chain reaction, a splicing isoform of hB1F (hB1F-2) was identified, which has an extra 46 amino acid residues in the A/B region. Examination of the tissue distribution has revealed an abundant 5.2-kilobase transcript of hB1F is present specifically in human pancreas and liver. Interestingly, an additional transcript of 3.8 kilobases was found to be present in hepatoma cells HepG2. Fluorescent in situ hybridization has mapped the gene locus of hB1F to the region q31-32.1 of human chromosome 1. Altogether, this study provides the first report that a novel human homolog of FTZ-F1 binds and regulates ENII of HBV. The potential roles of this FTZ-F1 homolog in tissue-specific gene regulation, in embryonic development, as well as in liver carcinogenesis are discussed.

Virus-induced hepatocarcinogenesis involves a series of histological developmental processes with the stepwise acquisition of several genetic changes that are necessary for the malignant transformation of hepatocytes. Although genetic alterations are known to be involved in the pathogenesis of hepatocellular carcinoma (HCC), little is known about the contributions of specific genes to this process. To gain insight into the genetic alterations involved in the neoplastic evolution from chronic hepatitis B virus infection to dysplastic nodules (DN) to HCC, we captured and sequenced the exomes of four DNA samples: one DN sample, two HCC samples, and one control peripheral blood sample from a single HCC patient. Mutations in the UBE3C gene (encoding ubiquitin ligase E3C) were observed in both tumor tissues. Then we resequenced the UBE3C gene in a cohort of 105 HCC patients and identified mutations in 17 out of a total of 106 (16.0%) HCC patients. The subsequent experiments showed that UBE3C promoted HCC progression by regulating HCC cells epithelial-mesenchymal transition. Clinically, a tissue microarray study of a cohort containing 323 HCC patients revealed that the overexpression of UBE3C in primary HCC tissues correlated with decreased survival (hazard ratio [HR] = 1.657, 95% confidence interval [CI] = 1.220-2.251, P = 0.001) and early tumor recurrence (HR = 1.653, 95% CI = 1.227-2.228, P = 0.001) in postoperative HCC patients. Conclusion: Our findings indicate that UBE3C is a candidate oncogene involved in tumor development and progression and therefore a potential therapeutic target in applicable HCC patients. (Hepatology 2014;59:2216–2227)

The Fc receptor associated pathway might improve the immune responses against hepatitis B virus (HBV) as previously described by us. In addition, the Flt3 ligand (FL) has been reported to potentiate antigen presenting cells in vivo and may act as a potential adjuvant to boost antigen-specific immune responses. In this study, the immune efficacies of a set of fusion proteins of HBsAg and Fc and/or FL were evaluated in HBsAg transgenic mice.The fusion proteins composed of HBsAg and the Fc domain of murine IgG1 (HBsAg-Fc) and/or the Flt3 ligand, and yeast-derived recombinant HBsAg were used as immunogen to immunize HBsAg transgenic mice, respectively. Serum and liver HBsAg levels, serum anti-HBsAg and cytokine profile, and the activities of alanine aminotransferase (ALT)/AST were investigated after immunization.After six injections, the most pronounced decrease in serum and liver HBsAg levels was observed in the HBsAg-Fc immunized group. In addition, serum Th1 cytokines and ALT/AST activities were highest in this group, indicating an effective induction of a favorable cellular immune response. Interestingly, the fusion protein containing HBsAg-Fc and the Flt3 ligand stimulated an alternative Th1-type immune response featured with high level productions of tumor necrosis factor α (TNF-α) and monocyte chemoabstractant protein 1 (MCP-1), causing a more severe cytotoxicity in hepatocytes while showed less effective in reducing serum HBsAg level.HBsAg-Fc is effective in eliciting both the humoral and cellular immune responses against HBsAg in HBsAg transgenic mice, which makes it a potential immunogen for the immunotherapy of chronic hepatitis B.

To clone Streptococcus salivarius (Ss) 57. I urease gene, which can express ureolytic activity in Escherichia coli (Ec) without adding extra nickel ions.Urease gene was cloned by polymerase chain reaction in three separate parts. The three separate plasmids were digested by specific restriction enzymes and ligated together. The expression of the complete urease gene in Ec was detected by phenol red assay and pH analysis.Urease gene of Ss 57.I was eventually cloned and proved correct. Urease activity of the obtained clone was positive in Ec. Without adding extra NiCl(2), the recombinant Ec could hydrolyze urea to produce ammonia, resulting in the increase of pH value.The clone of Ss urease gene obtained in this study could express ureolytic activity in Ec without adding extra nickel ions. The current clone can be used to construct ureolytic effector strain used in replacement therapy in caries prevention.

The genome of hepatitis B virus (HBV) consists of four open reading frames, encoding the envelope proteins (Pre-S/S), the core proteins (Pre-C/C), the polymerase (P) and the transactivating X protein (X). In the sera of HBV-infected patients, hepatitis B surface antigen (HBsAg) particles without the viral genome can outnumber virions by more than 1000-fold. To analyse the interactions between HBsAg and host cells, global gene-expression profiles of a small HBsAg (SHBs)-secreting stable cell line (HepG2-S-G2) and its counterpart control cell line (HepG2-Neo-F4) were compared. Marked upregulation of lymphoid enhancer-binding factor 1 (LEF-1), a transcription factor in the Wnt pathway, was found in SHBs-expressing cells and was confirmed by interference experiments with small interfering RNA. However, compared with the control cells, HepG2-S-G2 did not show higher proliferative competence in culture or increased tumorigenesis in nude mice. A possible mechanism to explain the discrepancy between the upregulation of LEF-1 and the lack of increased tumorigenesis is SHBs expression resulting in altered expression and distribution of LEF-1 protein in cell compartments and upregulation of LEF-1 isoforms that could suppress, rather than enhance, the Wnt pathway.

Human nuclear receptor hb1 f(nr5a2) was cloned and characterized as a novel member of the Ftz-F1 (nr5a) nuclear receptor subfamily,whose its biological function remained largely unidentified. The aim of this study was to establish transgenic mouse model that specifically expressed hB1F in the liver to faciliate the functional study of hB1F. hb1f cDNA was placed downstream of mouse albumin gene enhancer/promoter to construct a liver-specific hb1f expression vector. Transgene fragments were microinjected into fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. Four offspring were identified as carrying the transgenes by PCR,from which one was also verified by Southern blotting. RT-PCR and Western blotting results showed that the transgene was expressed in the liver of the transgenic mice. Transgenic founder mice were used to establish transgenic mouse lineages. The F1 mice were identified by PCR analysis. Genetic analysis of the transgenic mice demonstrated that the transgene had been integrated into the chromosome at a single site and could be stably transmitted.

Enhancer II (ENII) is one of the critical cis-elements in the Hepatitis B Virus (HBV) genome for the hepatic viral gene transcription and DNA replication. The liver-specific activity of ENII is regulated by multiple liver-enriched transcription factors, including LRH-1/hB1F, HNF1, HNF3β, HNF4 and C/EBP. Knowledge on the interplay of these important factors is still limited. In this study, we demonstrate a functional synergism between the orphan nuclear receptor LRH-1/hB1F and the homeoprotein HNF1 in up-regulating the liver-specific activity of ENII. This synergism is sufficient for initiating the viral gene transcription and DNA replication in non-hepatic cells. We have defined the activation domains in hB1F and HNF1 that contribute to the synergism. We further show that hB1F and HNF1 can interact directly in vitro and have mapped the domains required for this interaction.

Human hepatitis B virus enhancer II B1 binding factor (hB1F also known as NR5A2, LRH-1, FTF or CPF) is an orphan nuclear receptor and belongs to the fushi tarazu factor I (FTZ-F1) subfamily. It plays important roles in the transcriptional regulation of a number of genes involved in bile acid biosynthesis pathway, hepatitis B virus (HBV) replication and liver specific regulatory network. Like other nuclear receptors, hB1F is composed of modular functional domains. We characterized a domain in its hinge region that imposes a strong repression on the transcriptional activity of hB1F, which is important for the function of hB1F on regulating the activity of HBV enhancer II/core promoter. Mutations of the core residues in this domain abrogate the repression. Bioinformatic analysis reveals that the amino acid sequence of this region is highly conserved only among members of the FTZ-F1 subfamily. The repression is observed in five cell lines tested, while the degree of the repression varies greatly, which does not parallel with the expression level of the DEAD box protein of 130 kD (DP103), a potential interacting protein of a homologous domain in the steroidogenic factor 1 (SF-1). Moreover, the repression is not affected by the silencing mediator for retinoic acid receptor and thyroid hormone receptor (SMRT) and steroid receptor coactivator 1 (SRC-1). Collectively, these data suggest a novel regulatory mechanism for the transcriptional activity of hB1F.

The orphan nuclear receptor hB1F (also known as NR5A2, LRH-1, FTF or CPF) plays important roles in regulating the expression of several cellular and viral genes actively involved in a wide range of biological processes such as the bile acid biosynthesis, liver specific gene regulatory network and hepatitis B virus replication. The activity of nuclear receptors is regulated by multiple mechanisms, including coactivation and corepression. In this study, it was found that the silencing mediator for retinoic acid receptor and thyroid hormone receptor (SMRT) specifically represses the transcriptional activity of hB1F, on either GAL4 dependent reporter system or the hB1F-responsive HBV enhancer II/core promoter. The repression imposed by SMRT is observed in different cell lines. Interestingly, hB1F couldn t interact with SMRT directly, as demonstrated by mammalian two-hybrid analysis or GST pull-down assay. Taken together, it can be concluded for the first time that the transcriptional activity of hB1F is regulated specifically by the corepressor SMRT via an indirect mechanism.

A novel coronavirus (SARS-coronavirus, SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS) recently. The first step in coronavirus infection is binding of the viral spike protein to certain receptor on host cells. The spike protein is the main surface antigen of the coronavirus and there should be antibodies against spike protein in patients serum. Thus, to develop and expression protein fragment from spike protein gene are the purposes of this experiment. Partial spike gene fragments (751-1925 bp, 2005-3410 bp, 1-1925 bp and 32-3659 bp) and its intact gene were cloned into pET32 or pGEX vectors, and transformed into competent Escherichia coli BL21(DE3) (pLysS), respectively. 63, 78, 98, 160 and 164 kD fusion proteins were successfully expressed with amounts of 35%, 34%, 24%, 17% and 5% of total cell protein. The soluble parts of the cell crude extract were then partially purified by GST affinity chromatography.

MVA is a genetically modified vaccinia virus strain, which is replication defective and extremely safe for research and clinical use. To further improve the safety of MVA and achieve more efficient selection of recombinant constructs, a transient selection marker system has been developed, which contains vaccinia virus k1l gene flanked by two identical fragments. In this report, four recombinant MVAs were constructed with this system, and homologous recombination of recombinant MVAs was studied during the construction procedure. The results showed that the recombination frequency of these rMVAs was significantly high, though lower than that observed in other vaccinia virus strains. The k1l free rMVAs could be conveniently obtained after 3 or 4 blind passages. Our data indicated that recombinant MVA with a transient selection marker system was safe for use as live vaccine and gene therapy vector for human. In addition, blind passages could enhance the efficiency in isolation of k1l free recombinants

Human nuclear receptor hB1F is a novel member of the fushi tarazu factor I subfamily of nuclear receptor superfamily. The studies about its homologous genes indicate that hB1F may play a key role in regulating the metabolic homeostasis of cholesterol. After obtaining the founder mice carrying the transgene of hb1f by microinjection, each founder was mated to normal C57 mouse and the positive F1 by PCR identification of the same founder were intercrossed within sisters and brothers to establish the transgenic mouse lineage. The results of F1, F2 and offspring of test cross identification showed that the widely expressed hb1f transgenic mouse lineage was established successfully in this study. The tissue morphology of the transgenic lineage was also analyzed preliminarily.

AIM: To analyze the tissue morphologic phenotype and liver gene expression profile of hB1F transgenic mice. METHODS: Transgene expression was analyzed with RTPCR and Western blotting. For one of the transgenic mouse lines, tissue expression pattern of the transgene was also examined with immunochemical methods. Pathological analysis was used to examine the tissue morphologic phenotype of established transgenic mice. The liver gene expression profile of transgenic mice was analyzed with microchip, and some of the differentially expressed genes were verified with RT-PCR. RESULTS: The expressions of hB1F were shown in livers from 6 of 7 transgenic mouse lines. The overexpression of hB1F transgene did not cause pathological changes. Expressions of three genes were up-regulated, while down-regulation was observed for 25 genes. CONCLUSION: The overexpression of hB1F transgene may cause changes of gene expression profiles in the liver of transgenic mice.

AIM: Human hepatitis B virus enhancer 11131 binding factor (hB1F) was cloned and characterized as a novel member of the Ftz-F1 (NR5A) nuclear receptor subfamily. Although progresses have recently been made, its biological function remains largely unidentified. The aim of this study was to establish an hB1F transgenic mouse model to promote the functional study of hB1F. METHODS: Transgene fragments were microinjected into fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. The offsprings were identified by PCR and Southern blot analysis. Transgene expression was analyzed with RT-PCR and Western blot analysis. Transgenic founder mice were used to establish transgenic mouse lineages. The F1 and F2 mice were identified by PCR analysis. RESULTS: Seven mice were identified as carrying copies of transgene. RT-PCR and Western blotting results showed that the transgene was expressed in heart, liver, lung, kidney and stomach in one of the transgenic mouse lineages. Genetic analysis of the transgenic mice demonstrated that the transgene was integrated into the chromosome at a single site, and was transmitted stably. CONCLUSION: In this study we established an hB1F transgenic mouse model, which will facilitate the investigation of the biological function of hB1F in vivo.

The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) is an important viral structural protein. Based on bioinformatics analysis, 10 antigenic peptides derived from the S protein sequence were selected and synthesized. The antigenicity and immunoreactivity of all the peptides were tested in vivo and in vitro. Four peptides (P6, P8, P9 and P10) which contain B cell epitopes of the S protein were identified, and P8 peptide was confirmed in vivo to have a potential in serological diagnosis. By using a syncytia formation model, we tested the neutralization ability of all 10 peptides and their corresponding antibodies. It is interesting to find that P8 and P9 peptides inhibited syncytia formation, suggesting that the P8 and P9 spanning regions may provide a good target for anti-SARS-CoV drug design. Our data suggest that we have identified peptides derived from the S protein of SARS-CoV, which are useful for SARS treatment and diagnosis.