猪源马链球菌兽疫亚种表达的类M蛋白黏附抑制性单抗的获得

根据马链球菌兽疫亚种(Streptococcus equi subsp.zooepidemicus)猪源株ATCC35246株的类M蛋白基因序列,通过PCR技术,扩增出无信号肽的类M蛋白基因并定向克隆至表达载体pET-32a( )中。重组质粒经酶切鉴定和测序后,在大肠杆菌BL21中表达,获得60kDa产物,Western blot显示,其与ATCC35246株多克隆抗血清反应,抗原性良好。以His亲和层析柱纯化重组类M蛋白作为抗原免疫8周龄的BALB/c小鼠,采用淋巴细胞杂交瘤技术制备了12株稳定分泌抗类M蛋白单抗的细胞株,特异性检测显示其与A群链球菌、猪链球菌2型以及马链球菌马亚种等没有交叉反应。单抗亚类鉴定显示,其中6株为IgG1,3株为IgG2,另外3株为IgM。从腹水中纯化的单抗效价为2.56×104~1.01×105。黏附抑制实验显示,其中一株单抗能阻断类M蛋白黏附HEp-2细胞。     

Induction of an effective anti-tumor immune response and tumor regression by combined administration of IL-18 and Apoptin

Immunization strategies using plasmid DNA can potentially improve humoral and cellular immune responses that protect against cancer and infectious diseases. The chicken anemia virus-derived Apoptin protein exhibits remarkable specificity in its ability to induce apoptosis in tumor cells, but not in normal diploid cells. Interleukin-18 (IL-18) is a Th1-type cytokine that has demonstrated potential as a biological adjuvant in murine tumor models. In this study, we analyzed the anti-tumor potential and mechanism of action of simultaneous Apoptin and IL-18 gene transfer in C57BL/6 mice bearing Lewis lung carcinoma (LLC). Here we report that the growth of established tumors in mice immunized with pAPOPTIN in conjunction with pIL-18 was significantly inhibited compared with the growth of tumors in mice immunized with the empty vector (EV) or pAPOPTIN alone. Furthermore, the immunization of mice with pAPOPTIN in conjunction with pIL-18 elicited strong natural killer activity and LLC tumor-specific cytotoxic T lymphocyte (CTL) responses in vitro. In addition, T cells from lymph nodes of mice vaccinated with pIL-18 or pAPOPTIN + pIL-18 secreted high levels of the Th1 cytokine IL-2 and IFN-γ, indicating that the regression of tumor cells is related to a Th1-type dominant immune response. These results demonstrate that vaccination with Apoptin together with IL-18 may be a novel and powerful strategy for cancer immunotherapy.     

豆科植物中鱼藤酮含量测定及光学异构体研究

采用HPLC法测定了豆科鱼藤属的毛鱼藤、白花鱼藤、边荚鱼藤、马来鱼藤,灰豆属的非洲山毛豆、灰毛豆,豆薯属的豆薯、鸡血藤属厚果鸡血藤和紫穗槐属紫穗槐9种植物中鱼藤酮含量,发现毛鱼藤含量最高,每100g植物干粉中含1．82g鱼藤酮,紫穗槐含量最低,仅含0．005g。通过比较毛鱼藤、白花鱼藤、边荚鱼藤、非洲山毛豆、豆薯分离得到鱼藤酮的比旋光度,结合马来鱼藤、灰毛豆、厚果鸡血藤、紫穗槐提取物对3龄家蚕的点滴触杀活性差异,说明这些植物中含有的鱼藤酮均为同-光学异构体。     

Construct synthetic gene encoding artificial spider dragline silk protein and its expression in milk of transgenic mice

Based on the known partial cDNA sequence of dragline silk protein an artificial gene monomer, a 360 bp sequence, was designed and polymerized to encode an analog of dragline silk protein. Six tandem copies of monomer were cloned into pBC1 vector and microinjected into the pronuclei of fertilized Kunming White eggs. Transgenic mice were screened by Polymerase Chain Reaction (PCR) and Southern blot which revealed that 10 mice (5 male, 5 female) among 58 mice were transgenic positive. Milk of five F0 mice and eight F1 mice was analyzed by Western blot, and two F0 mice and seven F1 mice expressed recombinant dragline silk protein. In transgenic mice milk a maximum of concentration of recombinant dragline silk protein was 11.7 mg/L by radioimmunoassay.     

Small interfering RNA-mediated silencing induces target-dependent assembly of GW/P bodies

Gene silencing using small interfering RNA (siRNA) is a valuable laboratory tool and a promising approach to therapeutics for a variety of human diseases. Recently, RNA interference (RNAi) has been linked to cytoplasmic GW bodies (GWB). However, the correlation between RNAi and the formation of GWB, also known as mammalian processing bodies, remains unclear. In this report, we show that transfection of functional siRNA induced larger and greater numbers of GWB. This siRNA-induced increase of GWB depended on the endogenous expression of the target mRNA. Knockdown of GW182 or Ago2 demonstrated that the siRNA-induced increase of GWB required these two proteins and correlated with RNAi. Furthermore, knockdown of rck/p54 or LSm1 did not prevent the reassembly of GWB that were induced by and correlated with siRNA-mediated RNA silencing. We propose that RNAi is a key regulatory mechanism for the assembly of GWB, and in some cases, GWB may serve as markers for RNAi in mammalian cells.     

模拟降雨变化对古尔班通古特沙漠土壤养分及酶活性的影响

土壤酶参与土壤系统的养分循环过程,是联系植物和土壤养分的关键纽带。土壤酶活性对降水格局变化响应敏感,这种响应对于缺水且养分贫瘠的荒漠生态系统显得尤为重要。然而,早春积雪完全融化后首次降雨时间及降雨量如何影响土壤养分及土壤酶活性还鲜见相关报道。以新疆古尔班通古特沙漠为研究区,在早春积雪完全融化后,设置3个首次降雨时间(积雪完全融化后第10天、20天和30天)和3个降雨梯度(5 mm、10 mm和15 mm),于植物生长旺季采集土壤样品,研究土壤养分含量和土壤酶活性的响应特征。结果表明：积雪完全融化后不同首次降雨时间下5mm降雨处理以及积雪完全融化后第30天下各降雨量处理对土壤养分和酶活性影响不显著。积雪完全融化后第10天,随降雨量增加,土壤全碳呈显著先下降后增加趋势,全钾呈显著增加趋势,而土壤微生物量碳呈显著降低趋势;积雪完全融化后第20天,随降雨量增加,速效氮、土壤蔗糖酶活性、土壤微生物量碳氮呈先下降后增加趋势,土壤全碳和多酚氧化酶活性显著下降,土壤全钾和碱性磷酸酶活性显著增加。模拟10 mm降雨,随首次降雨时间推迟,土壤全氮、速效氮、速效磷、土壤蔗糖酶活性和土壤微生物量碳呈增加趋势;...     

基于MaxEnt模型的人类干扰对滇金丝猴潜在分布的影响

滇金丝猴 (Rhinopithecus bieti) 是我国特有的濒危灵长类动物,预测其栖息地变化,评估人类活动影响,进而识别保护空缺,对该物种的保护具有重要意义。本文使用出行大数据估计人类出行密度,并将这一指标引入栖息地模型的构建;运用MaxEnt模型和空间分析技术分别构建自然环境和人类干扰两种情景下滇金丝猴的适宜栖息地模型,并分析自然环境与人类干扰两类共计11个变量对栖息地的影响。结果显示：(1) MaxEnt模型的预测精度较高,基于出行大数据的人类出行密度这一指标能很好表征人类干扰对栖息地的影响。(2) 模型预测得到滇金丝猴高适宜栖息地面积3 487.28 km²,认为影响滇金丝猴潜在分布的主要环境变量是海拔、年降水量、人类出行密度和距道路距离。(3) 人类干扰对滇金丝猴栖息地有明显的负影响 (使适宜栖息地面积相较于自然环境下减少9.32%),其中人类出行活动对滇金丝猴的栖息地干扰最为明显;同时还发现研究区78.8%的区域均受到一个或多个人类变量的较强干扰。(4) 在现有15个滇金丝猴群的活动斑块中,3个同时具有较高栖息地适宜性和较多人类干扰的斑块,可作为重点保护区域;同时发现栖息地质量在距道路和居民点约2 500 m处均出现明显拐点,可作为开展保护工作的缓冲区参考距离。降低这些区域的人类干扰强度,对滇金丝猴的生存具有更重要的现实保护意义。