Expression and mechanisms of hsa_circ_087631 in patients with primary biliary cholangitis

AIM To explore the expression and mechanisms of circular RNA hsa_circ_087631 in the patients with primary biliary cholangitis （PBC）. METHODS RT-qPCR was used to detect the expression of hsa_circ_087631 in PBC patients and healthy controls. Hsa-miR-346-overexpressing lentiviral vector pLenti-EF1a-EGFP-F2A-Puro-CMV-MCS was constructed and transfected into human acute T cell leukemia Jurkat cells， and then the expression levels of hsa_circ_087631， Bcl-6 mRNA and interleukin-21 （IL-21） mRNA were detected by RT-qPCR. Dual-luciferase reporter assay was performed to identify the interactions between hsa_circ_087631 and hsa-miR-346. RESULTS The expression of hsa_circ_087631 in the PBC patients was significantly increased compared with the healthy subjects. Hsa-miR-346-overexpressing lentiviral vector pLenti-EF1a-EGFP-F2A-Puro-CMV-MCS was successfully constructed. The expression of hsa-miR-346 was significantly increased after the hsa-miR-346-overexpressing lentiviral vector was transfected into the Jurkat cells， while the expression levels of hsa_circ_0087631， Bcl-6 mRNA and IL-21 mRNA were significantly decreased. After wild-type or mutant hsa_circ_087631 vector and hsa-miR-346 mimics were transfected into 293T cells， the results of dual-luciferase reporter assay showed that hsa-miR-346 significantly decreased the luciferase activity of wild-type hsa_circ_087631 （P<0.01）， but the regulation did not change significantly after mutation of the predicted binding site. CONCLUSION Peripheral blood hsa_circ_087631 level is elevated in the PBC patients. The hsa_circ_087631/hsa-miR-346/Bcl-6 signaling may take effect in human T cells. Hsa-miR-346 significantly reduces the expression of hsa_circ_087631， but it may not be regulated by predicted binding sites.     

AM真菌对桑树根围土壤团聚体的影响机制

为揭示丛枝菌根 (Arbuscular mycorrhizal，AM)真菌对植桑土壤的影响及机制，采用盆栽试验研究接种摩西管柄囊霉 (Funneliformis mosseae，Fm)和根内根生囊霉 (Rhizophagus intraradices，Ri)对土壤有机碳(Soil organic carbon, SOC)、球囊霉素相关土壤蛋白 (Glomalin related soil protein, GRSP)及团聚体组成与稳定性的影响。结果表明：⑴ 接种Ri显著增加土壤大团聚体百分比，并提高平均质量直径 (Mean weight diameter, MWD)和几何平均直径 (Geometric mean diameter, GMD)、显著降低团聚体破坏率 (Percentage of aggregate destruction, PAD)。⑵ 接种Fm和Ri均显著增加微团聚体SOC含量，接种Fm显著降低大团聚体总GRSP含量，而接种Ri却显著增加大团聚体和微团聚体总GRSP含量及易提取GRSP含量。⑶ 接种AM真菌对整体SOC的效应为负，土壤总GRSP对SOC占比在25.5%~76.5%之间，土壤易提取GRSP对SOC占比在4.87%~5.93%之间，且Ri的接种效应高于Fm。⑷ 总GRSP、易提取GRSP和SOC对团聚体组成表现均为正向显著影响，其中易提取GRSP是主要驱动因子，而总GRSP是土壤团聚体稳定性的主要影响因子。综上，AM真菌作用下桑树根围土壤团聚体得以改善并趋于稳定，Ri的接种效应明显大于Fm；土壤团聚体的形成主要依赖易提取GRSP，而其稳定性主要受总GRSP影响。     

Nitrogen fertilization affects yields and storage compound contents in seeds of field-grown soybeans cv Enrei (Glycine max. L) and its super-nodulating mutant En-b0-1 through changing N2 fixation activity of the plants

ABSTRACT

We compared the protein and oil contents, seed yields, and natural abundance of ¹⁵N (σ¹⁵N) of seeds from the plants of the cultivar Enrei, which has moderate nodulating ability (Enrei) with those of its two near-isogenic lines (NILs), a super-nodulating genotype of Enrei, i.e., En-b0-1, and a non-nodulating genotype of Enrei, i.e., En1282. Plants of these three genotypes were grown on four types of field plots with different types of urea coated slow-release nitrogen (N) fertilizers (CUSLNFs) which delivered N in different manners during plant growth . The seed yield of the En1282 plants was similar to that of the Enrei plants but much greater than that of the En-b0-1 plants when the plants were grown on the field to which a CUSLNF with a long lifespan was applied. The amounts of protein per seed were positively correlated with those of oil per seed in the case of En1282 plants irrespective of the field N conditions. The seed protein contents were proportional to the seed weight in both En1282 and Enrei plants. Such a relationship was not observed in the En-b0-1 plants, and the protein and oil contents in the seeds of En-b0-1 plants were negatively correlated with each other. These observations suggested that the N supply to maturing seeds was a key to the protein contents in the En1282 and Enrei plants and that the C supply to maturing seeds was a key to the protein contents in the En-b0-1 plants. The comparison of the σ¹⁵N values, protein contents, and seed yields of seeds from En1282 plants suggested that the Enrei plants assimilated considerable amounts of N from the soil during the late plant growth. We concluded that high N₂ fixation activity depressed the amounts of oil per seed and changed the protein and oil contents in soybean seeds.     

狂犬病病毒HEP-Flury M基因重排后在小鼠神经母细胞瘤细胞中的表型分析

【目的】探究狂犬病病毒HEP-Flury M基因重排对基因转录和蛋白表达的影响,揭示病毒在小鼠神经母细胞瘤(NA)细胞中的表型变化与M基因重排的相关性。【方法】通过荧光定量PCR、Western blot以及病毒在NA细胞中的生长和扩散试验,对亲本毒株rHEP-Flury和M基因重排毒株M2、M4在NA细胞中的基因转录、表达、生长和扩散进行比较。【结果】狂犬病病毒结构基因的转录和表达主要受病毒基因组RNA合成的影响,但是在一个完整转录过程中单个结构基因的转录比例与其所在位置相关,M基因重排病毒的Leader RNA (LeRNA)和L mRNA的转录比例显著高于亲本毒株rHEP-Flury。M基因重排病毒在NA细胞中的生长和扩散都劣于亲本毒株rHEPFlury。【结论】狂犬病病毒亲本毒株rHEP-Flury具有狂犬病病毒原始的基因组顺序,在NA细胞中的生长和扩散都明显优于M基因重排病毒。结构基因在基因组中的位置主要决定其在一次转录过程中的转录比例,进而影响病毒在NA细胞中的生长和扩散。     

酸奶营养品质提升的加工工艺研究

牛奶中含有丰富的活性营养物质,包括乳铁蛋白、免疫球蛋白、乳过氧化物酶、α-乳白蛋白、β-乳球蛋白等。传统工艺生产的酸奶经过高温杀菌,活性营养物质几乎完全损失。为了避免这一情况,本文推出一种新的酸奶生产工艺,生产出的酸奶可保留大部分活性营养物质。首先将牛奶的乳脂进行分离,乳脂部分进行125 ℃、5 s杀菌,脱脂牛奶部分进行75 ℃、15 s杀菌,将两部分料液混合后加入菌种发酵至酸度75~85°T,破乳后冷却后熟8 h以上。经测定,新工艺酸奶中α-乳白蛋白、β-乳球蛋白、乳铁蛋白、免疫球蛋白、乳过氧化物酶含量分别为966 mg/L、1 963 mg/L、23.4 mg/L、102 mg/L、1 025 U/L,远高于传统工艺酸奶的68 mg/L、200 mg/L、0.7 mg/L、14 mg/L、0 U/L。通过对比新工艺与传统工艺酸奶保质期内的活菌数、黏度、酸度和口感,并无显著差异。     

拟南芥乙烯应答基因HLS1的原核表达及多克隆抗体制备

【目的】制备乙烯应答基因HLS1多克隆抗体,在过表达植物中检测HLS1的积累,为深入研究HLS1响应乙烯信号通路的分子机制奠定基础。【方法】构建pET28a-HLS1c原核表达载体,转化到大肠杆菌BL21(DE3)中,用异丙基-β-D-硫代半乳糖苷(IPTG)诱导获得重组蛋白HLS1c-His;镍柱亲和纯化重组蛋白后免疫家兔获得抗HLS1血清,利用抗原亲和纯化抗血清获得高纯度的HLS1多克隆抗体;用HLS1抗体和GFP抗体,检测原生质体瞬时转化体系中GFP-HLS1c融合蛋白的表达;用农杆菌蘸花法获得过表达GFP-HLS1g的转基因植物,用HLS1抗体和GFP抗体检测GFP-HLS1g融合蛋白的表达。【结果】成功构建了原核表达载体pET28a-HLS1c,并在大肠杆菌系统中诱导获得重组蛋白HLS1c-His,且多以包涵体形式存在。纯化的HLS1c-His多克隆抗体,能清晰检测到约80 ng原核表达的HLS1抗原。纯化的HLS1抗体不仅能检测到原生质体中瞬时表达的GFP-HLS1c融合蛋白,也能检测到过表达转基因植物株系中的GFP-HLS1g融合蛋白,并且与标签蛋白GFP对应抗体所检测到的特异性条带大小一致。【结论】成功制备了拟南芥HLS1蛋白多克隆抗体,为HLS1蛋白在植物发育中的功能研究奠定了基础。     

异源表达玉米ZmC3H54基因增强水稻的耐旱性

CCCH锌指蛋白是一类重要的转录调控因子。从玉米中分离得到一个受干旱和ABA诱导表达的CCCH型锌指蛋白基因ZmC3H54，通过构建过量表达载体并转化水稻来进一步研究其功能。 与对照组相比，转基因植株在干旱胁迫处理下具有更高的相对含水量与存活率以及较低的相对电导率，表明过量表达ZmC3H54基因可以提高转基因水稻的耐旱性。此外，转基因水稻幼苗对外源ABA敏感性更高。以上结果表明玉米ZmC3H54基因可能是通过ABA信号通路调控植物对干旱的耐受性。     

不同饲粮粗蛋白质水平下黑水虻蛋白替代豆粕对蛋鸡生产性能、蛋清品质及血清蛋白质代谢指标的影响

本试验旨在探讨不同饲粮粗蛋白质水平下黑水虻蛋白替代豆粕对蛋鸡生产性能、蛋清品质及血清蛋白质代谢指标的影响。采用单因素随机试验设计,选用252只产蛋率相近的33周龄罗曼白蛋鸡,随机分为3组,每组7个重复,每重复12只鸡。各组分别饲喂标准回肠可消化氨基酸(SIDAA)平衡模式下配制的不同粗蛋白质水平(16.50%、14.85%、13.20%)的玉米-豆粕-黑水虻蛋白饲粮,各组黑水虻蛋白与豆粕提供等量的粗蛋白质。预试期2周,正试期12周。结果表明:1)与16.50%组相比,14.85组%和13.20%组的产蛋率和平均日采食量无显著影响(P>0.05),平均蛋重显著降低(P<0.05),料蛋比显著升高(P<0.05);13.20%组的产蛋量显著降低(P<0.05),14.85%组的产蛋量无显著差异(P>0.05)。2)与16.50%组相比,14.85%组的平均蛋重、蛋清重、浓蛋白重、蛋白高度、哈氏单位和蛋清比例均无显著差异(P>0.05);13.20%组的平均蛋重、浓蛋白重、蛋白高度显著降低(P<0.05),蛋清重和哈氏单位呈下降趋势(P=0.0513和P=0.0673)。3)各组血清谷草转氨酶、谷丙转氨酶活性及总蛋白、白蛋白、尿酸含量无显著差异(P>0.05)。16.50%和13.20%饲粮粗蛋白质水平对蛋鸡输卵管膨大部分泌功能有轻微不良影响。由此可见,在SIDAA模式下,以黑水虻蛋白替代豆粕并使饲粮粗蛋白质水平降低至14.85%时对鸡蛋蛋清品质无不良影响。     

玉米瘤黑粉菌SG200效应蛋白PEP1表达及生物信息学分析

运用生物信息学手段分析pep1基因结构,并根据GenBank中玉米瘤黑粉菌pep1 DNA序列设计引物,从玉米瘤黑粉菌SG200的基因组中扩增得到了pep1基因全长,构建了pET–28a–pep1重组表达质粒,选用大肠埃希菌BL21(DE3)作为宿主菌,以1 mmol/L IPTG诱导表达。SDS–PAGE检测结果表明,诱导表达产物大小与理论值(20 800)一致,说明pET–28a–pep1能够在大肠埃希菌BL21(DE3)中高效表达。     

微生物发酵在菜籽饼粕饲用品质改良中的应用研究进展

随着畜牧业的快速发展、优质饲料原料的严重不足以及国际进口贸易形势的日益紧张,开发非常规饲料资源的需求愈发迫切。菜籽饼粕是我国第一大油料作物油菜的加工副产物,饲用价值不高严重制约了在饲料工业中的应用。微生物发酵技术具有改良菜籽饼粕品质和提高畜禽生产性能的优势。本文综述了微生物发酵改良菜籽饼粕饲用价值的机理、影响因素以及微生物发酵菜籽饼粕饲料在畜禽生产中的应用,讨论了目前微生物发酵菜籽饼粕生产中存在的问题,展望了未来微生物发酵菜籽饼粕饲料的发展方向。