转Bt基因大米暴露对雄性子代生殖系统影响的研究

目的 研究转Bt基因大米(TT51)暴露对Wistar大鼠雄性子代生殖系统发育的影响.方法 亲代雌雄大鼠分别连续给予市售大米、亲本大米和TT51大米70 d后,交配并产生子代,孕期和哺乳期各组雌性大鼠继续给予相应受试大米;子代雄性大鼠断乳后各组均给予普通饲料至70日龄,其间每周称量体重并记录食物消耗量和动物生长发育状况.雄性仔鼠至70日龄处死,进行血常规、血生化、血清性激素水平、生殖器官重量以及精子参数等指标检测.结果 TT51大米组与市售和亲本大米组比较,雄性子代体重、食物利用率、血常规、血生化、血清性激素水平以及精子各项指标差异无统计学意义.结论 转Bt基因大米暴露对雄性大鼠子代生殖系统发育未见不良影响.     

转Bt基因大米暴露对亲代雌性大鼠免疫系统影响的研究

目的 研究转Bt基因大米(TT51)对亲代雌性大鼠免疫系统的影响.方法 试验分为转Bt基因大米组(TT51组)、亲本明恢63大米组(明恢63组)和市售大米组,各组大米按60％的比例掺入饲料进行全食物喂养.Wistar雌、雄大鼠喂养10周后进行交配繁殖,对亲代雌性大鼠于断乳后进行免疫毒性评价.指标包括血常规、全血淋巴细胞分型、脾细胞的NK活性检测、刀豆蛋白A诱导淋巴细胞转化试验、抗体生成细胞检测以及淋巴结的淋巴细胞分型.结果 脾淋巴细胞分型中,TT51组的Th细胞、T细胞比例与市售大米组比较差异有统计学意义(P＜0.01),明恢63组的B细胞、T细胞比例与市售大米组比较差异有统计学意义(P＜0.01);颈部淋巴结淋巴细胞分型中,TT51组的Th细胞比例与市售大米组比较差异有统计学意义(P＜0.01);各组其他功能性试验结果之间差异无统计学意义.结论 转Bt基因大米(TT51)对亲代雌性大鼠免疫系统功能未见不良影响.     

大鼠繁殖试验评价牛初乳对生殖发育的影响

目的通过繁殖试验观察牛初乳对亲代大鼠生殖及F1代仔鼠发育状况的影响,评价牛初乳的生殖发育安全性。方法选用4周龄SD大鼠,牛初乳组在交配前期、宫内暴露期、哺乳期和仔鼠性成熟期(受孕前11周至仔鼠性成熟),用含10%牛初乳的饲料喂饲亲代和子代大鼠,对照组饲喂普通饲料,检测亲代体重、摄食量、动情周期、繁殖力指数、怀孕指数、孕周、每窝活产数、激素水平、生殖器官及精子质量;子代出生体重及身长、性别比、4日生存率、阴道开口时间、包皮分离时间等。采用两独立样本t检验的统计方法对均数进行比较,使用SPSS 13.0统计分析软件处理数据。结果与对照组相比,牛初乳对亲代的动情周期、繁殖力指数、怀孕指数、孕周、每窝活产数、子代性别比、出生身长、4日生存率均无显著影响,但亲代雄鼠左侧储精囊脏体比降低,精子总数提高(P<0.05),且牛初乳组亲代雌、雄大鼠血液中的促黄体生成素(LH)显著降低(P<0.05),雌鼠血液中催乳素(PRL)和孕酮(P)均显著低于对照组(P<0.05)。结论一代繁殖试验未发现牛初乳对生殖发育能力有显著影响,仅见亲代储精囊、精子总数和激素水平改变,其长远影响需二代繁殖试验进一步验证。     

早期给予膳食槲皮素对大鼠母代生殖及其子代生长发育的影响

目的:研究早期给予槲皮素对母鼠生殖及其子代发育的影响。方法:孕鼠随机分为4组:空白对照组、槲皮素低、中、高剂量组(从孕0d至断乳21d灌胃给予槲皮素,剂量分别为16、160、480mg/(kg.d))。定期检测母鼠和仔鼠的生长指标、繁殖能力、幼仔存活及幼仔肛殖距(AGD),产后第21天处死母鼠和仔鼠,计算母鼠和幼仔脏器指数。结果:与空白对照组相比,槲皮素组对母鼠体质量、繁殖能力、幼仔存活率没有影响;槲皮素低、中剂量组中雄性仔鼠AGD和体质量显著降低;槲皮素组对雌性仔鼠AGD和体质量没有影响;槲皮素高剂量组降低母鼠和雄性仔鼠的脾脏指数,中剂量组增加雌性仔鼠肾脏指数。结论:槲皮素具有雌激素及抗雌激素作用,对雄性仔鼠影响较大,无生殖与发育毒性。     

两代繁殖实验研究牛初乳粉对大鼠生殖发育的影响

为探讨围产期接受牛初乳粉对子代生殖发育的影响,设计了两代繁殖实验,分析亲代和子代雌雄鼠血清激素水平,生殖器官发育和生殖受孕变化。结果表明：食用含有牛初乳粉的饲料后,亲代和子代雌鼠生殖受孕指标及雄鼠精子密度和活率与空白组相比无显著差异（p〉0.05）。亲代和子代雄鼠血清激素水平与空白组相比无显著差异（p〉0.05）,但亲代雌鼠血清中催乳素和孕酮显著低于空白组（p〈0.05）,子一代血清中催乳素显著低于空白组（p〈0.05）。子二代雌鼠F2a生殖器间距与空白组相比显著降低（p〈0.05）,子二代F2b的阴道开口时间推迟（p〈0.05）,不能确定是偶然发生还是受生理影响所致,需要重复实验的验证。     

杭菊花乙醇提取物对大鼠生殖发育毒性研究

目的研究杭菊花乙醇提取物(CME)经过亲代、宫内、哺乳期和断乳后持续暴露对子代的影响。方法设对照组和CME低、中和高剂量组,CME掺入饲料,剂量分别为0、1、3、9 g/kg BW;参照OECD TG 443开展一代延长试验,每组孕鼠20只及其子鼠每组40只(雌雄各半),子鼠断乳后持续给予CME 13周,测定生长发育、生殖发育、血象、血生化指标,做组织病理学检查。结果 CME高、中剂量组子鼠睁眼时间明显迟于对照组;低剂量组子代雌性大鼠出生后(PND)56～112天体质量明显高于对照组,子代雌性大鼠CME高、低剂量组红细胞计数(RBC)和血红蛋白浓度(HGB)均降低,CME高剂量组谷丙转氨酶(ALT)升高,高、中剂量组谷草转氨酶(AST)、血磷(P)升高,CME各剂量组血糖(GLU)均降低、乳酸脱氢酶(LDH)均升高;子代雄性大鼠CME高、低剂量组血清钾(K~+)降低;子代雌雄大鼠CME高剂量组脾脏/体质量系数均高于对照组;以上均差异有统计学意义(F=2.818～42.75,P0.05)。结论低剂量CME促进子代雌性大鼠体质量生长发育,降低子代雌性大鼠的血糖,影响RBC、HGB和LDH;高、中剂量CME影响子代大鼠发育和子代雌性大鼠的肝功能,高剂量CME影响子代雌雄大鼠脾脏/体质量系数。     

槲皮素对肥胖母鼠后代生长发育的影响

目的：研究孕、哺乳期给予槲皮素对肥胖母鼠后代生理和神经发育的影响。方法：建立营养性肥胖雌性SD大鼠模型,交配受孕,随机分成5组,每组16只孕鼠,整个实验期进食高脂饲料,并分别以0、50、100、200mg/kg剂量的槲皮素灌胃;以16只正常体质量孕鼠作为空白对照,整个实验期喂食基础饲料并以溶剂灌胃。记录仔鼠出生体质量,并观察主要生理和神经发育指标。结果：高脂对照组后代出生体质量显著高于空白对照组,随槲皮素干预剂量增大,出生体质量逐渐降低,接近空白对照水平。该作用持续到断乳。生理发育：高脂对照组雄性仔鼠睾丸下降迟于空白对照组,槲皮素能在一定程度上逆转此影响,200mg/kg效果最显著;神经发育：200mg/kg组的平面翻正、悬崖回避反射达标时间明显早于空白对照组和高脂对照组。结论：肥胖孕鼠后代出生体质量显著增加,而孕期槲皮素干预可有效逆转新生仔鼠体质量,控制其哺乳期体质量过快增长。一定剂量的槲皮素能够有效逆转孕期肥胖造成的雄性后代睾丸下降延迟,并能够促进后代特定神经反射的形成。     

转Bt玉米暴露对亲代及子代大鼠免疫细胞分型及细胞因子的影响

为研究转苏云金芽胞杆菌(Bacillus thuringiensis,Bt)基因抗虫玉米长期食用对大鼠免疫的影响,以其亲本非转基因玉米饲料及商业饲料为对照,分别采用流式细胞术及液相芯片法检测喂养90 d后亲代及子代刚断乳大鼠外周血、脾脏免疫细胞分型及相关血清细胞因子的表达。结果发现,转Bt基因抗虫玉米饲料组亲代大鼠外周血和脾脏及子代脾脏各淋巴细胞亚群(CD4~+、CD8~+、TCRαβ~+、TCRγδ~+)的细胞数量比例及亲代血清中10种细胞因子表达水平均与非转基因玉米饲料组无显著差异(P0.05);子代外周血B淋巴细胞数量比例显著增高,巨噬细胞炎症蛋白-1α、单核细胞趋化因子-1β、白细胞介素-5表达水平明显降低(P0.05)。结果表明:转Bt基因抗虫玉米暴露90 d对亲代大鼠免疫系统影响不大;对子代刚断乳大鼠的细胞免疫及体液免疫有一定的影响。     

转GmDREB3抗旱基因小麦大鼠一代生殖发育毒性研究

目的研究转GmDREB3抗旱基因小麦对大鼠生殖发育的影响。方法无特定病原体(SPF级)Wistar大鼠随机分为基础饲料组、亲本对照组、转基因组,每组60只,雌雄各半,分别给予基础饲料、亲本对照饲料和转基因饲料,13周后组内雌雄大鼠交配产生F1代仔鼠,分析指标有:F0代和F1代大鼠的毒性表现;F0代大鼠的体质量、进食量、食物利用率;F0代雌鼠繁殖指数和性激素水平(促黄体生成素释放激素、卵泡刺激素、雌二醇);F0代雌雄大鼠脏器系数、肝脑比、脾脑比及生殖器官病理;F1代仔鼠体质量、身长、尾长、出生后21天神经病理(大脑、小脑和脑干)和出生后56天雌鼠免疫学指标(分泌性免疫球蛋白A、免疫球蛋白E、溶菌酶、二胺氧化酶)。结果与亲本对照组比较,F0代雄鼠肝脏系数降低(P0.01)、脾脏系数升高(P0.05)、脾脑比升高(P0.01),均差异有统计学意义;F1代仔鼠出生后第0天、第4天、第7天、第21天的尾长偏短,差异有统计学意义(P0.05),其他指标均差异无统计学意义(P0.05)。结论转GmDREB3抗旱基因小麦对大鼠生殖发育无毒性作用。     

亲代暴露致敏原对子代BN大鼠致敏模型的影响

研究BN大鼠亲代暴露致敏原后子代对致敏原免疫反应变化。方法 亲代分成暴露与非暴露致敏原,两组所产大鼠均随机分为卵清蛋白(OVA)组、空白对照组,分别每日经口给予1 mg/ml OVA溶液、灭菌蒸馏水1 ml,共6周;第2周、第4周、第6周取血分离血清,测定特异性抗体IgG和IgE;第3天和第5天取血分离血浆,测定组胺;最后测定两组胃肠道渗透性。结果 亲代非暴露致敏原的大鼠可激发较强的过敏反应,特异性IgG、特异性IgE、组胺水平均升高;亲代暴露致敏原的大鼠激发后免疫反应较弱。结论 BN大鼠亲代接触致敏原后,其子代致敏免疫反应减弱。     

Toxicological evaluation of transgenic rice flour with a synthetic cry1Ab gene from Bacillus thuringiensis

Bacillus thuringiensis (Bt) transgenic (KMD1) and non‐transgenic (KMD1′s parental variety Xiushui 11) rice flours were assessed in a 90 day feeding test with rats. KMD1 contained a synthetic cry1Ab gene from Bt, and selection marker genes nptII and hpt linked in tandem. In the≤64 g kg^?1 body weight (BW) dosage range (Bt transgenic rice flour composed 64% of the ingredients of the diet), no adverse effects of Bt rice on rats were observed in terms of animal behaviour, weight gain and feed utilisation rate. Necropsy at the end of the experiment indicated that neither pathological lesions nor histopathological abnormalities were present in organs such as liver, kidneys, intestines and testes of rats in both test and control groups by macroscopic and microscopic pathology. In addition, no significant differences (P > 0.05) were observed in relative organ weights, haemograms and blood indices of rats between test and control groups. Several serum parameters of female rats were found to be significantly different between Bt and non‐Bt diets, but the values of these parameters were still within the normal ranges of values for rats of this age and sex. These results demonstrated that Bt rice flour at a dosage of 64 g kg^?1 BW, Bt toxin and NPTII and HPT proteins have no toxic effects on rats. © 2002 Society of Chemical Industry     

转Bt基因水稻种子中Bt蛋白含量测定方法的研究

采用ELISA技术检测转Bt基因水稻中Bt蛋白的含量,判断水稻样品中是否含有转基因成分。利用研磨、酶标、孵育等技术对样品进行前处理。采用阳性质控物浓度等倍稀释方法,建立标准曲线,相关系数为0.997 4。用一系列不同转基因含量的标准基体材料,分析方法的最低检测限,灵敏度达0.1%。通过对我国进入生产性试验的转Bt基因水稻品系TT51-1和科丰6号的测试,表明该方法与普通PCR方法真实性和灵敏度一致,可以广泛应用于转Bt基因水稻及其粗加工产品的转基因成分检测。为转基因生物安全监管和安全性评价提供技术支撑。     

转基因水稻Bt63的外源基因相对含量分析

为分析转基因Bt63稻米外源基因含量,利用新型、灵敏和高通量的实时荧光定量PCR技术,以RBE-4作为转基因水稻的内源参照基因,通过梯度稀释法,分别获得了Bt 63和RBE-4基因的Ct值与起始模板相关性的标准曲线,相关系数分别为0.996 6和0.995 4。通过转基因水稻外源基因(Bt63)和内标基因(RBE-4)起始模板数的比较,测定了外源基因在转基因水稻中的相对含量,这项研究将为建立转基因水稻抗虫能力评价体系和转基因稻谷的流通和监管提供技术支撑。     

转人乳铁蛋白基因大米的慢性毒性研究

目的 观察转人乳铁蛋白(hLF)基因大米是否具有慢性毒性作用.方法 初断乳的180只SD大鼠按性别、体重随机分为3组:转基因hLF基因大米组、亲本大米对照组和AIN-93对照组,分别饲喂相应饲料12个月.观察大鼠的体重、进食量、血常规、血生化情况,实验末期处死动物,称量脏器重量并对脏器进行病理学检查.结果 转hLF基因大米组血常规(LYM％、GRN％)、血生化(ALT、AST、GLU)在个别时间点上与亲本对照组或AIN-93对照组存在显著差异(P＜0.05);其他观察指标与两个对照组均无显著差异.结论 现有实验结果不能证实转hLF基因大米对大鼠有慢性毒性作用.     

Detection of genetically modified rice: collaborative validation study of a construct-specific real-time PCR method for detection of transgenic Bt rice

A collaborative trial study has been conducted for validation of an extraction method and a subsequent real-time PCR for detection of a transgenic Bt rice line (‘Bt63’) in rice products originating from China. A total of 17 laboratories participated in the study and each laboratory received 16 coded samples comprising of rice grain flours, rice noodle flours and plasmid DNAs. Of the accepted results all Bt63-positive rice grain samples (0.1 or 0.05% w/w) and all rice noodle samples prepared from marketed rice products were detected correctly. The result demonstrates that ‘Bt63’ rice is detectable even at low relative mass concentrations of 0.05%. The absolute LOD determined with plasmid DNA samples showed to be at least five copies of the ‘Bt63’ target sequence. The data provided in this study show that the method is fit-for-purpose to inspect Chinese rice products for the presence of EU-unauthorised rice lines carrying the ‘Bt63’ construct.     

Real-time PCR method for detection of the transgenic rice event TT51-1

The insect-resistant transgenic rice event TT51-1 (synonym BT63) has been found illicitly planted and distributed for years although it has never been approved for commercial cultivation in any country up to now. The purpose of this study was to establish a detection method that is specific for this transformation event. The event-specific PCR method produces an amplicon of 120 basepairs (bp) based on the revealed 3′ junction sequence with a limit of detection (LOD) and a limit of quantification (LOQ) being approximately 5 and 10 initial template copies, respectively. Two mixed rice samples with known TT51-1 contents were used to verify the developed real-time PCR system, from which the expected results were observed.     

Detection of genetically modified rice: a construct-specific real-time PCR method based on DNA sequences from transgenic Bt rice

Genetically modified rice varieties developed in China are close to approval for agricultural cultivation and production. However, so far no method has been reported for specific detection of transgenic varieties of this crop. In the present study, rice seeds assumed to consist of field-tested Bt rice (‘Anti-pest Shanyou 63’ and ‘Anti-pest Jinyou 63’) were used as reference material to determine transgenic DNA sequences. The transition between the cryIA(b) and cryIA(c) fusion gene and the nopaline synthase terminator (nos) sequence was used to develop a construct-specific real-time PCR based detection method. This Bt rice specific detection system was combined with a recently published quantitative real-time PCR method for the rice-specific (Oryza sativa L.) reference gene gos9. The complete PCR assay for detection of transgenic Bt rice was in-house validated and the limit of quantification was found to be below 0.1% Bt rice relative to the rice content. Application of the PCR assay should allow more precise detection of transgenic rice varieties in imported food products which are so far not approved in the EU.