Sequence Analysis and Prokaryotic Expression of NSP1 Gene of Bovine Rotavirus UK Strain

Rotavirus is a major cause of acute diarrhea in both many kinds of young animals and children under 5 years old.Rotavirus NSP1, a 55 ku RNA binding protein, is the product of gene 5, which can subvert innate immune responses and be one of virulent determinant factors.According to the sequence in GenBank, specific primers targeting to NSP1 gene were designed and the gene was amplified by RT-PCR, following by being cloned into the pET-28a(+) vector.It showed that the full length of NSP1 gene was 1 473 bp, encoding 491 amino acids.The NSP1 shared the highest identity with WC3 strain.The recombinant protein was induced in E.coli Rosetta(DE3) by IPTG and was analyzed by SDS-PAGE and Western blotting.The results revealed that NSP1 recombinant protein existed in the form of inclusion body with the molecular weight of 55 ku.The purified recombinant protein could be recognized by His-tag antibody.This study laid the foundation for further research on the relationship between the intracytoplasmic location of NSP1 protein and its activity.     

Mycoplasma pneumoniae induces IL-1β production through activating NLRP3 inflammasome by ROS in RAW264.7 cells

AIM: To investigate whether Mycoplasma pneumoniae (Mp)-induced interleukin-1β (IL-1β) production in RAW264.7 cells is through the activation of NLRP3 inflammasome via reactive oxygen species (ROS). ME-THODS: RAW264.7 cells were randomly divided into 3 groups. In normal group, RAW264.7 cells were treated without Mp. In model group, RAW264.7 cells were treated with 1∶ 10 multiplicity of infection (MOI) of Mp. In NAC group, RAW264.7 cells were pretreated with N- acetylcysteine (NAC) at a concentration of 5 mmol/L for 30 min before infection with Mp. The RAW264.7cells were infected with Mp (1∶ 10 MOI) for 4, 8, 16 and 24 h in model group and NAC group, respectively. The intracellular ROS level was analyzed by flow cytometry. The mRNA expressions of NLRP3, ASC and caspase-1 were detected by real-time PCR. The protein levels of NLRP3, ASC and caspase-1 p20 were determined by Western blot. The levels of pro-inflammatory cytokine IL-1β in the supernatant were measured by ELISA. RESULTS: Compared with normal group, the production of ROS were significantly increased at 4, 8, 16 and 24 h after infection, the mRNA expression of NLRP3, ASC and caspase-1 were increased at 8, 16 and 24 h after infection, the protein levels of NLRP3, ASC and caspase-1 p20 were increased at 16 and 24 h after infection, and the releases of IL-1β were increased at 24 h after infection in model group (P<0.01). Compared with the model group, the level of ROS in NAC group decreased, so as the expression of NLRP3, ASC and caspase-1 at mRNA and protein levels and the releases of IL-1β in the supernatant at the corresponding time points. CONCLUSION: Mp may stimulate the ROS production to activate NLRP3 inflammasome in RAW264.7 cells.     

菜籽油裂解燃料离子液体催化酯化反应降酸工艺的研究

为了降低裂解燃料酸值,增加燃料的稳定性能,将吡啶丁烷磺酸硫酸氢盐应用于菜籽油裂解燃料的酯化反应。考察了催化剂用量、反应时间和反应温度等对酸值的影响,并在最佳优化条件下考察了裂解燃料成品的理化性质。结果表明:吡啶丁烷磺酸硫酸氢盐对催化酯化反应具有很高的催化活性。优化工艺条件为:催化剂用量1.2%、反应温度75℃、反应时间70 min,在此工艺条件下,酸值降低到1.0 mgKOH/g以下。     

中辽1号杨在辽宁地区引种研究

对20世纪末新引进到辽宁并经初选的12个杨树无性系进行了引种试验研究,通过无性系多点生长对比试验、室内外物候观测、对杨干象抗性评价和材性测试等,筛选出生长快、适应强的杨树新品种中辽1号杨,其材积生长量分别超过当地对照种辽宁杨、沙兰杨、荷兰3930杨和I-214杨14.8%~68.1%。其生长进程中高生长最快期是在5~6年生,连年生长量最大数值达到3.6 m;胸径生长最快期是在3~5年生,连年生长量最大数值达到6.34 cm。而且干型好、材质优良,对杨干象有相对较强的抗性,可作为新品种在其适生区大面积推广。     

Ultrastructural impairment of islet microvascular endothelial cells in STZ-induced type 1 diabetic mice

AIM: To investigate the ultrastructural changes of islet microvascular endothelial cells in STZ-induced type 1 diabetic mice. METHODS: BALB/c mice were randomly divided into diabetic group and control group. The expression of insulin and platelet-endothelial cell adhesion molecule-1 (CD31) in islet microvessels was detected by immunohistochemical staining. The ultrastructural changes of islet β cells and islet microvessels were observed under transmission electron microscope. RESULTS: Compared with control group, the number of islet β cells, ratio of β cells/α cells, average number of secretory granules in β cells and insulin expression area per islet in diabetic group were significantly decreased (P<0.01). Besides, diabetic group had fewer microvessels with lower expression of CD31 (P<0.01). Mitochondria in islet microvascular endothelial cells and pericytes in diabetic group were swelling. The basement membrane of islet microvessels became thicker in diabetic group (P<0.01). CONCLUSION: Islet microvascular endothelial cells were impaired in type 1 diabetic mice.     

Cloning and Sequence Analysis of Buffalo Keap1 Gene and Investigation of Its Expression Pattern in Different Tissues

In this study,the CDS sequence of buffalo Keap1 gene was cloned and analyzed,then its expression pattern in different tissues was also investigated.A pair of primers of buffalo Keap1 gene was designed based on the nucleotide sequence of Bos taurus Keap1 gene from GenBank,and then the buffalo Keap1 gene was amplified.Using the bioinformation techniques,the gene sequence and the protein structure were analyzed.The expression level of Keap1 gene in different tissues were detected with Real-time quantitative PCR.The results showed that the length of buffalo Keap1 gene coding sequence was 1 875 bp and encoded 624 amino acids.The multiple sequence alignment results showed that buffalo Keap1 gene shared 99%,96%,92% and 90% of similar nucleotide sequence with that of Bos taurus,Ovis aries,Sus scrofa and Homo sapiens,respectively.And the phyogenetic tree also showed the conservatism between several different species.The second structure of buffalo Keap1 protein was predicted as 24 alpha regions,40 beta regions,38 turn regions and 27 coil regions.In addition,the results of Real-time quantitative PCR showed that Keap1 mRNA exists in all seven tissues,but the most abundant expression was in heart and the minimal expression was in liver and spleen.The results provided an foundation for further study of Keap1-Nrf2-ARE signal pathway,for enhancing the ability of antioxidant of buffalo embryo in vitro culture.     

江汉平原棉花合理施氮量研究

采用田间小区试验研究了江汉平原棉花合理施氮量。结果表明,在施用90 kg/hm~2 P2O5、180 kg/hm~2K2O和3 kg/hm~2持力硼基础上,利用线性+平台模型得到棉花中高产量水平下的合理施氮量为280 kg/hm~2。在不明显减产的条件下,从提高氮肥当季利用率、尽量降低氮肥投入的角度,可以将氮肥用量降低到240kg/hm~2左右,在此施氮水平下仍然可能通过改进田间管理措施获得高产。因此,江汉平原棉花氮肥减量空间为20~60 kg/hm~2。     

Ss-Bi1 encodes a putative BAX inhibitor-1 protein that is required for full virulence of Sclerotinia sclerotiorum

Sclerotinia sclerotiorum is a destructive necrotrophic plant pathogen with global distribution. Although S. sclerotiorum has been studied extensively, substantial research on aspects of the pathogen's ability to cause disease is still needed. Bax inhibitor-1 protein functions as a suppressor of programmed cell death and is involved in the response to biotic and abiotic stress in animals, plants and yeast. In this study, we functionally characterized a putative Bax inhibitor-1 protein, Ss-Bi1, from S. sclerotiorum. Ss-Bi1 is predicted to contain a BAX inhibitor-1-like super family domain and shows significant homology with many BAX inhibitor-1 proteins. High expression levels of Ss-Bi1 were observed in hyphae under various stresses. Targeted silencing of Ss-Bi1 resulted in reduced virulence in host plants. Ss-Bi1 gene-silenced strains were more sensitive to heat stress and ER stress than the wild-type strain. The results suggest that Ss-Bi1 encodes a putative BAX inhibitor-1 protein that is required for full virulence of S. sclerotiorum.