斑马鱼g型溶菌酶基因序列分析及其原核表达

【目的】实现斑马鱼g型溶菌酶在大肠杆菌中表达,以获得高纯度且具溶菌活性的融合蛋白,为探究g型溶菌酶在斑马鱼抗菌过程中的作用机制及其开发利用打下基础。【方法】通过ClustalW、ExPASy、SignalP-5.0 Server及PSIPRED等在线软件对斑马鱼g型溶菌酶进行生物信息分析,经密码子优化后合成斑马鱼g型溶菌酶基因,亚克隆至pET-28a（+）表达载体并转化大肠杆菌BL21（DE3）感受态细胞,以IPTG进行诱导表达,并通过非干扰型蛋白浓度测定试剂盒和溶菌酶测定试剂盒（比浊法）测定其浓度及溶菌活性。【结果】从GenBank检索获得的斑马鱼溶菌酶基因g1（NM_001002706.1）和g2（XM_002664371.5）分别命名为Zeb-Lys-g1和Zeb-Lys-g2。Zeb-Lys-g1基因开放阅读框（ORF）长591 bp,共编码196个氨基酸残基,其编码蛋白分子量约21.6 kD;Zeb-Lys-g2的ORF长576 bp,共编码191个氨基酸残基,其编码蛋白分子量约21.1 kD;2种斑马鱼g型溶菌酶序列中均含有2个半胱氨酸残基及3个保守的催化残基位点（Glu、Asp和Asp）。Zeb-Lys-g1的N端含有17个氨基酸的信号肽,而Zeb-Lys-g2不存在典型的信号肽结构,但其三级结构均具有多个α-螺旋结构。在基于溶菌酶序列相似性构建的系统发育进化树中,Zeb-Lys-g1序列与金鱼g型溶菌酶序列的亲缘关系最近,而Zeb-Lys-g2序列与鲈形目和鲽形目鱼类的g型溶菌酶序列亲缘关系较近。将合成的斑马鱼g型溶菌酶基因（Zeb-Lys-g1和Zeb-Lys-g2）亚克隆至pET-28a（+）表达载体并转化BL21（DE3）感受态细胞,20 ℃下经IPTG（终浓度0.5 mmol/L）诱导16 h,可获得融合蛋白Zeb-Lys-g1和Zeb-Lys-g2,纯化后的浓度分别为1.01和1.66 mg/mL,对应的溶菌活性分别为689.68和44.39 U/mg。【结论】鱼类g型溶菌酶的基因变异较保守,经密码子优化及全基因合成方式合成的斑马鱼g型溶菌酶基因Zeb-Lys-g1和Zeb-Lys-g2可通过原核表达获得高纯度、高溶菌活性的融合蛋白,为探究斑马鱼溶菌酶的抗菌机制提供了技术支持。     

植物细胞程序性死亡研究进展

细胞程序性死亡(PCD)是植物体一定发育阶段普遍存在的一种现象,它受特定的基因调控,是主动的细胞死亡过程。介绍了PCD的概念、一般特征、机理、与植物衰老的差别以及检测技术等方面的研究进展。对植物PCD研究中有待进一步解决的问题和实际意义进行了展望。     

水产动物营养与免疫研究进展

随着水产养殖集约化与规模化的发展,水产品为人类提供了大量优质食物蛋白质,为国家粮食安全、解决我国“三农”问题做出了重要贡献。长期研究实践证明,营养与饲料科技贡献在水产养殖中占有更加重要的地位。饲料组成不仅影响水产动物的营养,而且会影响水产动物的免疫与抗病能力,营养状况是决定水产养殖动物抵抗疾病的能力的重要因素之一。特别是在水产集约化、规模化养殖中,水产养殖动物面临着大量的应激,如：营养、环境因子等激烈变化,易诱发疾病,甚至死亡。反之,水产养殖动物的生理与健康状况也影响饲料的利用与代谢。营养和免疫的关系显著地影响着水产养殖生产,一直是大家关注的热点。在过去的5年中,水产动物营养与免疫方面的研究取得了重大进展,对蛋白质(氨基酸)、脂肪、碳水化合、维生素、矿物元素等多种必需营养素及其免疫、抗病能力等方面做了大量的研究。本文就营养与免疫相关的研究进展做一综述,以供参考。     

Effects of DEHP on testosterone synthesis in fetal Leydig cells of newborn male rats

AIM: To observe the effects of diethylhexylphthalate(DEHP) on testosterone synthesis in fetal Leydig cells(FLC) of newborn male rats.METHODS: The pregnant rats were exposed to DEHP at dose of 10 mg·kg^-1·d^-1, 100 mg·kg^-1·d^-1 or 750 mg·kg^-1·d^-1(body weight) by gavage from gestation 12 days(GD 12) to postnatal 1 day(PND 1) respectively. The serum level of testosterone was detected by chemiluminescence. The morphology of FLC from the testes was observed under light microscope and transmission electron microscope. The mRNA expression of steroidogenic acute regulatory protein(StAR) and insulin-like growth factor I(IGF-I) was detected by real-time PCR(ΔΔCT). RESULTS: The serum testosterone level in low dose group was significantly higher than that in control, middle and high dose groups. The serum testosterone level in middle and high dose groups was significantly lower than that in control group(P<0.05). Light microscopy showed the aggregative cluster distribution of FLCs in low dose group, while manifested as tumor-like hyperplasia of FLCs in middile dose group and high dose group. Under electron microscope, the FLC in low dose group showed oval-shape or long spindle-shape, the lipid particles were decreased, but smooth endoplasmic reticulum and mitochondria were increased in cytoplasm. In middle and high dose group, the FLC were spindle or oval-shaped, showed large or small, nuclear large, round, rich in cytoplasm and cell aggregation, the cytoplasm was rich lipid particles with deep-stained, smooth endoplasmic reticulum and mitochondria were expanded. Compared to control group, the mRNA expression of StAR in low, middle and high dose group was decreased. The mRNA expression of StAR in high dose group was decreased more significantly(P<0.01) as compared to that in control group, while the mRNA expression of IGF-I in low dose group and middle dose group was increased, but that only in low dose group was increased more significantly as compared to control group(P<0.01). CONCLUSION: DEHP has toxic effect on FLC and changes the morphology and steroidogenic capacity in testicular FLC. The low dose of DEHP elevates the gene expression of IGF-I, and IGF-I stimulates the production of testosterone by FLC. The high dose of DEHP may inhibit the gene expression of StAR to reduce the serum levels of testosterone.     

茄子果实性状相关基因的QTL定位

以果实性状差异显著的茄子(Solanum melongena L.)种质材料绿茄‘131'和上海本地长茄‘141'为作图亲本,构建了237个株系的F_2群体,采用450对SSR标记构建遗传图谱,定位茄子果实单果质量、果长、果径、果形指数和果实花萼大小的QTL。研究结果表明:在亲本间表现多态性的SSR标记为63对,平均多态性比例为14%;共有53个标记被分配到13个连锁群上(12条染色体),13个连锁群覆盖总长度为532.2cM,标记间距范围为0.2~45.9 cM,平均距离为10.04 cM;共定位了9个与果实性状相关的QTL位点(单果质量1个,果长1个,果径2个,果形指数3个,果实花萼大小2个)。     

猪组织和血浆中磺胺二甲嘧啶残留分布规律的数学模型分析

采用高效液相色谱法检测猪肌肉、肾脏、肝脏和血浆中的磺胺二甲嘧啶(Sulfamethazinum,SM2)残留量.通过对饲料中SM2添加量和组织、血浆中残留量回归关系不同数学模型的分析,确立残留分布规律的最佳数学模型.饲料中SM2的添加量在100～300μg/g的范围内,饲料添加量与猪组织(肌肉、肾脏、肝脏)、血浆中的残留数学模型分别为:y=0.241 6 e0.0061x,y=1.969 1 e0.0025x,y=3.049 5 e0.0019x,y=4.386 2 e0.0037x.通过对猪活体血浆样品的检测,根据建立的最佳残留数学模型,可算出饲料中SM2的添加量和肌肉、肾脏、肝脏中SM2的残留量,实现活体的监测.     

皖西白鹅球虫种类及感染情况调查

采用传统的卵囊形态学方法,结合球虫寄生部位,对皖西白鹅球虫的感染率和种类进行了调查和鉴定。共查48个粪样,球虫阳性48份,阳性率100％。共检到1科2属6种球虫,其中艾美耳属（Eimeria）5种,即鹅艾美耳球虫（E．anseris）、棕黄艾美耳球虫（E．fulva）、赫曼艾美耳球虫（E．hermani）、有害艾美耳球虫（E．novcens）和多斑艾美耳球虫（E．stigmosa）;泰泽属（Tyzzeria）1种,即稍小泰泽球虫（T．parvula）。稍小泰泽球虫检出率最高（100％）,其次是赫曼艾美耳球虫（50．0％）,为本次调查的优势种。     

不同碳源促进污染水体氮素转化的微生态过程

在试验条件下,向氮污染河道水体中C/N按15∶1补加甲醇、乙醇、乙酸钠、柠檬酸钠、葡萄糖、蔗糖、麦芽糖、羧甲基纤维素钠,检测水体氨氮、亚硝酸盐、硝酸盐、总氮的变化,以及在此过程中水体细菌胞外酶活性、细菌群落结构的变化。结果显示,除甲醇、羧甲基纤维素钠外,补加有机碳源显著降低了水体氨氮的含量,水体β-葡萄糖苷酶活性在碳源补加后快速增加,补加葡萄糖、蔗糖、麦芽糖的水体中细菌群落结构明显异于其他处理,表明添加有机碳源可显著加快污染水体中氮素的转化,水体微生物在此过程中起到关键作用。水体细菌群落结构和代谢活性对不同类型有机碳源有着不同的响应,因而添加不同有机碳源对水体氮素转化具有不同的促进作用。     

Settling of insecticide from dip wash mixed with dam water and zinc sulphate and used to control sheep lice (Bovicola ovis)

SUMMARY Insecticidal dipping fluid emulsions, mixed in vitro in dam water containing suspended clay particles and 1% w/v zinc sulphate, were analysed to determine rates of settling of diazinon, cyhalothrin and cypermethrin. Fifteen minutes after mixing, the concentration of the insecticides 5 cm below the surface had declined by 72.5%, 72.8% and 89.4%, respectively. On remixing, the concentration of insecticide in suspension was close to or greater than the initial concentration. In 2 trials, lice were eradicated from sheep showered with dip wash mixed in cloudy dam water to which 1% w/v of zinc sulphate was added. In 12 flock treatments in which 1000 to 2000 sheep were dipped with added zinc sulphate, the concentration of insecticide remained above the minimum lethal for susceptible strains of lice. However, lice were still present 6 months later in 8 of these flocks. When zinc sulphate is added to dip wash, agitation is needed to maintain the insecticide in suspension.     

安徽部分地区鸡球虫对六种药物的抗药性研究

选择56只土杂鸡,检测了采自安徽部分地区的7种艾美耳球虫对6种常用抗球虫药的敏感性。根据最适抗球虫活性百分率、病变记分减少率和相对卵囊产量3项指标综合判定,安徽部分地区的柔嫩艾关耳球虫、毒害艾美耳球虫、巨型艾美耳球虫、堆型艾关耳球虫、和缓艾美耳球虫、早熟艾关耳球虫和哈氏艾美耳球虫的7个混合种对地克珠利、尼卡巴嗪、托曲珠利、磺胺喹嗯啉钠、磺胺氯吡嗪钠和盐霉素钠均存在不同程度的抗药性,其中对盐霉素钠、地克珠利和托曲珠利具有重度抗药性,对尼卡巴嗪、磺胺喹嗯啉钠和磺胺氯吡嗪钠具有中度抗药性。