H9N2禽流感病毒M2 DNA疫苗初免-蛋白加强免疫的保护效果研究

流感病毒M2(基质蛋白2)是A型流感病毒的一个高度保守的蛋白。由于其免疫原性较弱,本研究采用M2 DNA疫苗初免-蛋白加强的策略来考察M2的免疫保护效果。制备A/Chicken/Jiangsu/07/2002(H9N2)流感病毒的M2 DNA疫苗以及经大肠杆菌表达的去除M2跨膜区的M2蛋白即sM2。以SPF级BALB/c小鼠为模型,电击法免疫M2 DNA疫苗,滴鼻法免疫sM2蛋白,免疫间隔三周,并于末次免疫后三周以致死量5LD50流感病毒H9N2攻击小鼠,通过检测小鼠存活率、体重丢失率、肺部病毒滴度及IgG抗体水平等指标来评价免疫的保护效果。实验结果表明,基于M2的疫苗采用DNA疫苗初免蛋白加强免疫二次的免疫程序能诱导较高的特异性抗体,明显减轻小鼠流感病症,提供完全的保护。     

Mouse-rabbit germinal vesicle transfer reveals that factors regulating oocyte meiotic progression are not species-specific in mammals

A series of experiments were designed to evaluate the meiotic competence of mouse oocyte germinal vesicle (GV) in rabbit ooplasm. In experiment 1, an isolated mouse GV was transferred into rabbit GV-stage cytoplast by electrofusion. It was shown that 71.8% and 63.3% of the reconstructed oocytes completed the first meiosis as indicated by the first polar body (PB1) emission when cultured in M199 and M199 + PMSG, respectively. Chromosomal analysis showed that 75% of matured oocytes contained the normal 20 mouse chromosomes. When mouse spermatozoa were microinjected into the cytoplasm of oocytes matured in M199 + PMSG and M199, as many as 59.4% and 48% finished the second meiosis as revealed by the second polar body (PB2) emission and a few fertilized eggs developed to the eight-cell stage. In experiment 2, a mouse GV was transferred into rabbit MII-stage cytoplast. Only 13.0-14.3% of the reconstructed oocytes underwent germinal vesicle breakdown (GVBD) and none proceeded past the MI stage. When two mouse GVs were transferred into an enucleated rabbit oocyte, only 8.7% went through GVBD. In experiment 3, a whole zona-free mouse GV oocyte was fused with a rabbit MII cytoplast. The GVBD rates were increased to 51.2% and 49.4% when cultured in M199 + PMSG and M199, respectively, but none reached the MII stage. In experiment 4, a mouse GV was transferred into a partial cytoplasm-removed rabbit MII oocyte in which the second meiotic apparatus was still present. GVBD occurred in nearly all the reconstructed oocytes when one or two GVs were transferred and two or three metaphase plates were observed in ooplasm after culturing in M199 + PMSG for 8 hr. These data suggest that cytoplasmic factors regulating the progression of the first and the second meioses are not species-specific in mammalian oocytes and that these factors are located in the meiotic apparatus and/or its surrounding cytoplasm at MII stage.     

四种白蚁的线粒体DNA限制性片段长度多态性研究(等翅目:鼻白蚁科,木白蚁科)

用６种限制性内切酶分析了４种白蚁的线粒体 Ｄ Ｎ Ａ 限制性片段长度多态性,根据限制性片段差异计算了４种白蚁之间的遗传距离,利用 Ｕ Ｐ Ｇ Ｍ Ａ 聚类分析法构建了分子聚类图。结果表明：尖唇异白蚁与散白蚁属关系很近,聚类分析结果显示应将其归于散白蚊属     

长江源区同域分布兔狲、藏狐和赤狐的时空重叠度

兔狲(Otocolobus manul)、藏狐(Vulpes ferrilata)和赤狐(V. vulpes)是青藏高原三江源区域分布的重要小型食肉兽。本研究于2014年6月至2019年9月在青海省长江源区沱沱河和通天河沿岸选取208个位点布设红外相机, 通过所获取的时空分布数据比较了上述3种同域分布小型食肉兽的时空利用情况。通过空间重叠度系数的比较分析, 兔狲和藏狐、兔狲和赤狐以及藏狐和赤狐之间的空间重叠度系数分别为0.25、0.48和0.17, 这表明兔狲、藏狐和赤狐三者在空间利用上存在一定的差异。通过核密度估计方法分析, 兔狲和藏狐属典型的昼行性动物, 而赤狐以夜行性活动为主。兔狲、藏狐和赤狐每个物种在冷暖两季的日活动节律重叠指数分别为0.83、0.78和0.88。两两比较分析表明, 兔狲和藏狐二者的日活动节律重叠指数最高(0.84), 兔狲和赤狐在夜间活动时段存在一定重叠(0.63), 而藏狐和赤狐的时间生态位分化最明显, 重叠指数最低(0.48)。此外, 在暖季, 两两物种之间的日活动节律重叠指数均小于其冷季的重叠指数。综上所述, 长江源区兔狲、藏狐和赤狐3种小型食肉兽可通过空间和时间资源的利用差异来降低物种间的干扰和竞争, 从而达到同域物种共存的目的。     

CD3ε和PMA对fas基因转录调控作用的研究

研究了ＣＤ３ε和ＰＭＡ对细胞凋亡基因ｆａｓ的转录调控作用．根据已知的人ｆａｓ基因５＇上游序列设计引物,用ＰＣＲ法从人胸腺细胞基因组ＤＮＡ中扩增出ｆａｓ５＇上游长为４４６ｂｐ的启动子片段．将此片段定向克隆到以虫荧光素酶为报告基因的真核细胞表达载体ｐＸＰ２中．经限制性内切酶酶切鉴定及序列测定分析表明此重组表达质粒ＤＮＡ的结构和序列正确．用此质粒（ｐＸＰ２－ｆａｓｕｐ）瞬时转染人ＪｕｒｋａｔＴ淋巴细胞,分析报告基因的表达水平．结果表明ｆａｓ基因上游－５０６到－６０的区域有弱启动子活性,约是阴性对照的１．５倍．抗ＣＤ３ε抗体和ＰＭＡ处理均可增强ｆａｓ启动子的活性,促进报告基因的表达,其虫荧光素酶活性分别是未经处理的ｐＸＰ２－ｆａｓｕｐ转染细胞的１．７倍和３．３倍,但二者没有协同作用．ＰＫＣ的抑制剂ｓｔａｕｒｏｓｐｏｒｉｎｅ和ＰＴＫ的抑制剂ｈｅｒｂｉｍｙｃｉｎＡ可抑制ＰＭＡ诱导的ｆａｓ基因转录,使虫荧光素酶基因的表达降至未用ＰＭＡ处理的水平．但ＰＴＫ的另一种抑制剂ｇｅｎｉｓｔｅｉｎ可与ＰＭＡ发挥协同作用,上调ｆａｓ基因的转录,使虫荧光素酶活性增加到５倍．这些结果为阐明ＣＤ３ε及ＰＭＡ介导Ｔ淋巴细胞激活和凋亡的分子机制     

北部湾沙田水域中华白海豚和江豚的同域分布格局及时空变化

北部湾是中华白海豚和江豚的重要分布区,但是对该水域两物种的分布格局及时空变化的研究却比较少,也不清楚这两个物种分布区是否有重叠。2003 ～ 2004 年与2011 ～2012 年期间,我们在北部湾沙田水域进行了船只调查,总计航程4 876.8 km,收集了两个物种空间分布的数据。结果表明,2003 ～ 2004 年仅发现中华白海豚,2011 ～2012 年则同时发现了中华白海豚和江豚。与2003 ～ 2004 年4 ～ 5 月相比,中华白海豚在2011 年4 ～ 5 月份的遇见率明显增加(0.152 vs 0. 028 头次/km)。中华白海豚的MCP、95% FK 家域和50% FK 核心域面积分别为162. 6 km² 、271. 35 km² 和56.78 km² ,2011 ～ 2012 年50% FK 核心家域有所扩大且东移。江豚的MCP、95% FK家域和50% FK 核心域面积分别为97.21 km² 、318.72 km² 和38.18 km² 。江豚主要分布于中华白海豚的外围远岸水域,同时两个物种的分布区有小部分重叠。基于多种因素的分析,我们推测,江豚可能存在季节性迁移以及中华白海豚对河口区有明显的偏好,可能是造成它们分布格局不同的原因。     

棉铃虫发生与北太平洋海温的遥相关 及其长期灾变预警

本文分析了山东郓城26年（1974～1999）、德州22年（1978～1999）和江苏丰县20年（1980～1999）棉铃虫百株累计卵量与北太平洋海温的遥相关关系及其时空动态规律,并选出相关显著程度P<0.05概率水平、空间分布范围较大、持续时间较长而稳定的组合作为关键预测因子组建了郓城、德州棉铃虫三代卵,丰县棉铃虫二代卵的预测模型,并筛选出最优长期灾变预警模型。结果表明：① 北太平洋海温场与棉铃虫种群数量消长存在显著或极显著的遥相关区域,其位置及范围随时间变化,但存在若干呈现出空间稳定性和时间持续性的大面积相关显著区域。② 郓城棉铃虫三代卵量和丰县棉铃虫二代卵量与北太平洋海温场的相关区分布形式很相似,与前两年1月份北太平洋月平均海温场存在大片相关显著的区域（35°～ 55°N,135°E～135°W）,持续时间达4个月之久;而德州棉铃虫三代卵量与前两年7～9月份北太平洋低纬度海温有大范围相关显著区（1°～17°N,165°E～120°W）。 ③ 用前两年1～11月份北太平洋海温场相关显著区内各格点的月平均海温距平的平均值做因子建立了棉铃虫长期灾变预警模型,预测检验结果表明：郓城棉铃虫三代卵6年（1994～1999）中报准5年,丰县棉铃虫二代卵5年（1995～1999）中报准3年,德州棉铃虫三代卵5年（1995～1999）全部符合。据此可提前20～27个月做棉铃虫的长期灾变预警。     

鸡白细胞介素2增强传染性法氏囊病病毒多聚蛋白DNA疫苗免疫原性的研究

鸡白细胞介素 2(IL-2)基因是新近被确定的非哺乳类IL-2基因。将鸡白细胞介素2(IL-2)基因和传染性法氏囊病病毒 (IBDV)多聚蛋白基因 (VP2/VP4/VP3)分别插入真核表达载体pCI的CMV启动子下游 ,制备DNA疫苗 ,免疫 14日龄SPF鸡 ,14d后二免 ,二免后 3d攻击标准强毒株。结果表明共注射鸡IL 2质粒能明显增强DNA疫苗对强毒攻击 ,保护率达 80 % ;能增强DNA疫苗诱导的中和抗体效价 (P<0.05 ) ;能显著促进鸡胸腺、脾脏和外周血液T淋巴细胞及法氏囊B淋巴细胞增殖反应(P<0.05)。这些结果提示鸡IL 2能明显增强IBDV多聚蛋白DNA疫苗的免疫原性 ,是一种优良的禽类DNA疫苗佐剂。     

6—DMAP和胚胎干细胞代数对异种重构卵体外发育的影响（简报）

The development of reconstructed oocytes and the survival rate of cloned animal were affected by many factors during nuclear transfer. The genetic constitution and the genetic state of donor nucleus were proposed to be primary factors, which affected the survival rate of cloned animal. In addition, the survival rate of cloned animal might be influenced by nuclear transfer technique itself and passages of donor cells as well as the activation methods of oocytes. We reconstructed oocytes with outbreeding Kunming albino mouse ES cells and enucleated rabbit oocytes, and analyzed the effects of the passages of ES cells and 6-DMAP on the development of interspecific reconstructed oocytes. The interspecific reconstructed ES-rabbit oocytes were activated either by combined two set electric pulses and 6-DMAP or by two set electric pulses alone. The rate of cleavage was significantly higher for the group (86.2%) treated with 6-DMAP than the group (64.2%, P < 0.05) treated with electric pulses only, and the rate of blastocysts was 17.0% and 13.4% respectively, which were not significantly different between two groups. When ES cells that had been passed for 24 and 14 generations were used as donors, the cleavage rates of the reconstructed oocytes were 88.5% and 82.1%, respectively (P > 0.05), and the rates of blastolation were 16.7% and 15.4%, respectively (P > 0.05). The results show that 6-DMAP increases the cleavage rate of reconstructed oocytes derived from ES cells, and affects slightly the developmental rate of blastocysts. There are no differences when high passage and low passage ES cells are used as nuclear donors.     

高产申嗪霉素和吩嗪-1-酰胺的水稻根际铜绿假单胞菌PA1201分离、鉴定与应用潜力

【目的】从水稻根际筛选能高效抑制水稻病原菌的细菌,分析和鉴定其形态和生化特征,为开发新型绿色农药奠定基础。【方法】从水稻根际分离能以1-氨基环丙烷-1-羧酸(ACC)为唯一碳源的菌株,根据菌株形态和生化特性、16S r DNA序列比对和磷脂脂肪酸图谱,对该菌株进行鉴定。通过氯仿萃取抽提、高效液相色谱分析,确定菌株PA1201在PPM培养基和黄豆粉培养基中申嗪霉素和吩嗪-1-酰胺的产量。【结果】菌株PA1201能有效抑制水稻纹枯病菌和水稻白叶枯病菌的生长,属于铜绿假单胞菌(Pseudomonas aeruginosa sp.PA1201);PA1201产生两种抑菌代谢产物申嗪霉素和吩嗪-1-酰胺,在PPM和黄豆粉培养基中申嗪霉素的产量可达81.7 mg/L和926.9 mg/L,吩嗪-1-酰胺的产量亦可达18.1 mg/L和489.5 mg/L;PA1201产生大量胞外蛋白酶,对人肺腺癌细胞系A549和黑腹果蝇具有一定毒性。【结论】PA1201的申嗪霉素产量比现有生产菌株的出发菌株M18高3-4倍,还能产生另一种抑菌活性更高的衍生物吩嗪-1-酰胺,具有进一步开发的潜力。