18%. Compared to hFGF21, the biostabilities of rHSA-hFGF21, including their resistance to temperature and trypsinization were significantly enhanced, and its plasma half-life was extended by about 27.6 times. Moreover, the fusion protein rHSA-hFGF21 at medium and high concentration showed a better ability to promote glucose uptake after 24 h of stimulation in vitro. In vivo animal studies showed that rHSA-hFGF21 exhibited a better long-term hypoglycemic effect than hFGF21 in type 2 diabetic mice. Our results demonstrated a small-scale production of rHSA-hFGF21, which is important for large-scale production and clinical application in the future.Long non-coding RNA (lncRNA) has become an important regulator of many cellular processes, including cell proliferation. Although studies have shown that a variety of lncRNAs play an important role in the occurrence and development of hematopoietic malignancies, a more comprehensive and unbiased method to study the function of lncRNAs in leukemia cell lines is lacking. Here, we used short hairpin RNA (shRNA) library combined with high-throughput sequencing to screen lncRNAs that may affect the proliferation of leukemia cell lines, and identified lncRNA C20orf204-203 among 74 candidate lncRNAs in this study. Further experiments showed that C20orf204-203 was localized in the cytoplasm in both K562 and THP-1 cell lines. C20orf204-203 knockdown decreased the proliferation of K562 and THP-1 cell lines accompanied with the increased proportion of early apoptotic cells. We observed the increased mRNA level of BAD gene while decreased protein level of TP53 and BCL2. The expression of Caspase 3 decreased and Caspase 3-cleaved protein increased in THP-1 cell line. However, their changes were inconsistent in the two cell lines. Our experimental results showed that knockdown of lncRNA C20orf204-203 in leukemia cell lines affected cell proliferation although the mechanism of action in different cell lines may differ. Importantly, our research demonstrated the feasibility of using shRNA library combined with high-throughput sequencing to study the role of lncRNA in leukemia cell lines on a large scale.Influenza B virus (IBV) is more likely to cause complications than influenza A virus (IAV) and even causes higher disease burden than IAV in a certain season, but IBV has received less attention. In order to analyze the genetic evolution characteristics of the clinical strain IBV (B/Guangxi-Jiangzhou/1352/2018), we constructed genetic evolution trees and analyzed the homology and different amino acids of hemagglutinin and neuraminidase referring to the vaccine strains recommended by World Health Organization (WHO). https://www.selleckchem.com/products/10074-g5.html We found that strain B/Guangxi-Jiangzhou/1352/2018 was free of interlineage reassortment and poorly matched with the vaccine strain B/Colorado/06/2017 of the same year. We also determined the median lethal dose (LD50) and the pathogenicity of strain B/Guangxi-Jiangzhou/1352/2018 in mice. The results showed that the LD50 was 105.9 TCID50 (median tissue culture infective dose), the IBV titer in the lungs reached peak 1 d post infection and the mRNA level of the most of inflammatory cytokines in the lungs reached peak 12 h post infection. The alveoli in the lungs were severely damaged and a large number of inflammatory cells were infiltrated post infection. The study demonstrated that the clinical strain IBV (B/Guangxi-Jiangzhou/1352/2018) could infect mice and induce typical lung inflammation. This will facilitate the research on the pathogenesis and transmission mechanism of IBV, and provide an ideal animal model for evaluation of new vaccines, antiviral and anti-inflammatory drug.In this study, we aimed to construct a non-replication mRNA platform and explore the side effects of electroporation-mediated delivery of mRNA on the mice as well as the expression features of the mRNA. With luciferase gene as a marker, in vitro transcription with T7 RNA polymerase was carried out for the synthesis of luciferase-expressed mRNA, followed by enzymatic capping and tailing. The mRNA was delivered in vivo by electroporation via an in vivo gene delivery system, and the expression intensity and duration of luciferase in mice were observed via an in vivo imaging system. The results demonstrated that the mRNA transcripts were successfully expressed both in vitro and in vivo. The electroporation-mediated delivery of mRNA had no obvious side effects on the mice. Luciferase was expressed successfully in all the mRNA-transduced mice, while the expression intensity and duration varied among individuals. Overall, the expression level peaked on the first day after electroporation and rapidly declined on the fourth day. This study is of great importance for the construction of non-replication mRNAs and their application in vaccine or antitumor drug development.Breast cancer is the most common tumor in female, which seriously threatens the health of women. Triple-negative breast cancer is a subtype with the worst prognosis because of its special physiological characteristics and lack of targeted drugs. Therefore, it is urgent to develop new targeted treatments to improve the prognosis and survival rate of the patients. Previous studies have shown that heat shock protein gp96 is expressed on the membrane of a variety of cancer cells but not on the normal cells. Cell membrane gp96 levels are closely related to the poor prognosis of breast cancer, which may serve as a new target for breast cancer treatment. Based on the structure of gp96, we designed an α-helical peptide p37 that specifically targeting the ATP binding region of gp96. To improve the stability and decrease the degradation of the peptide, the N-terminus or C-terminus of p37 was coupled to PEG2000 or PEG5000 respectively, and four PEGylated polypeptides were obtained mPEG2000CY, mPEG5000CY, mPEG2000LC, and mPEG5000LC. The PEGylated polypeptides inhibited the proliferation and invasion of breast cancer cell SK-BR-3, among which mPEG2000CY showed the most significant inhibitory effect. The half-life of mPEG2000CY in vivo was significantly longer than p37, and it effectively inhibited the growth of xenografted tumors of triple-negative breast cancer MDA-MB-231. The results provide a basis for the development of new targeted drugs against breast cancer, especially the triple-negative breast cancer.A fusion protein containing a tetanus toxin peptide, a tuftsin peptide and a SARS-CoV-2S protein receptor-binding domain (RBD) was prepared to investigate the effect of intramolecular adjuvant on humoral and cellular immunity of RBD protein. The tetanus toxin peptide, tuftsin peptide and S protein RBD region were connected by a flexible polypeptide, and a recombinant vector was constructed after codon optimization. The recombinant S-TT-tuftsin protein was prepared by prokaryotic expression and purification. BALB/c mice were immunized after mixed with aluminum adjuvant, and the humoral and cellular immune effects were evaluated. The recombinant S-TT-tuftsin protein was expressed as an inclusion body, and was purified by ion exchange chromatography and renaturated by gradient dialysis. The renaturated protein was identified by Dot blotting and reacted with serum of descendants immunized with SARS-CoV-2 subunit vaccine. The results showed that the antibody level reached a plateau after 35 days of immunization, and the serum antibody ELISA titer of mice immunized with recombinant protein containing intramolecular adjuvant was up to 166 240, which was significantly higher than that of mice immunized with S-RBD protein (P less then 0.05). At the same time, the recombinant protein containing intramolecular adjuvant stimulated mice to produce a stronger lymphocyte proliferation ability. The stimulation index was 4.71±0.15, which was significantly different from that of the S-RBD protein (1.83±0.09) (P less then 0.000 1). Intramolecular adjuvant tetanus toxin peptide and tuftsin peptide significantly enhanced the humoral and cellular immune effect of the SARS-CoV-2 S protein RBD domain, which provideda theoretical basis for the development of subunit vaccines for SARS-CoV-2 and other viruses.Zinc transporter 8 (ZnT8) is an important candidate antigen for type Ⅰ diabetes. The autoantibody detection kit based on ZnT8 can be used to help diagnose type Ⅰ diabetes, and the related products have been launched in Europe and the United States. Since the recombinant production system of active ZnT8 has not been established in China, this key raw material is heavily dependent on imports. We used Saccharomyces cerevisiae to carry out the recombinant expression of ZnT8. First, multiple antigenic forms of ZnT8 were designed as C-terminal haploid (C), C-terminal diploid (C-C), and N-terminal and C-terminal concatemers (N-C). The proteins were expressed, purified and tested for antigenicity by bridging-type ELISA. The serum of 13 patients with type Ⅰ diabetes and the serum of 16 healthy volunteers were detected. C, N-C, and C-C proteins had similar detection rates, which were 53.8% (7/13), 61.5% (8/13) and 53.8% (7/13). The specificity of the three groups was 100% (16/16). The detection value on positive samples P3, P4, and P8 increased by more than 90%, indicating better serum antibody recognition ability. Finally, N-C protein was selected for further serum sample testing, and the test results were characterized by receiver operating characteristic (ROC) curve for sensitivity and specificity. Compared with imported gold standard antigen, the sensitivity was 76.9% (10/13) and the specificity was 87.5% (14/16). There was no significant difference in the sensitivity of the method, but the specificity needed to be improved. In conclusion, the ZnT8 N-terminal and C-terminal concatemer protein developed based on S. cerevisiae expression system is expected to be a key alternative raw material in the development of in vitro diagnostic reagents for type Ⅰ diabetes.This paper aims to explore the effects of chicken interferon-γ (ChIFN-γ) and interleukin-2 (ChIL-2) on type 1 helper (Th1) T lymphocyte differentiation. To be specific, ChIFN-γ and ChIL-2 were first expressed in Escherichia coli competent cells and then purified by Ni-NTA affinity chromatography. Different concentration of ChIFN-γ and ChIL-2 were employed to stimulate the lymphocytes in chicken peripheral blood which had been activated by concanavalin A (Con A), and the mRNA levels of cytokines related to Th1 cell differentiation were detected by real-time quantitative PCR (RT-qPCR). The results showed that both ChIFN-γ and ChIL-2 can significantly up-regulate mRNA levels of cytokines related to Th1 cell differentiation and the optimal concentration was 12.5 μg/mL and 25.0 μg/mL, respectively. In addition, specific-pathogen-free (SPF) chickens were immunized with ChIL-2 or ChIFN-γ together with H9N2 vaccine, or H9N2 vaccine alone by oral administration or intramuscular injection, respectively. The mRNA levels of cytokines related to Th1 cell differentiation were detected after immunization. The results showed that ChIFN-γ and ChIL-2 significantly up-regulated the mRNA levels of cytokines related to Th1 cell differentiation induced by H9N2 vaccine compared with H9N2 vaccine alone, and that the intramuscular injection was better than oral administration. In this study, we verified that ChIFN-γ and ChIL-2 can significantly enhance mRNA levels of cytokines related to Th1 cell differentiation induced by ConA or H9N2 vaccine in vitro and in vivo. The results of this study can lay a theoretical basis for using ChIFN-γ and ChIL-2 as vaccine adjuvants.