为快速检测家畜衣原体,选取家畜衣原体ompA基因的高度保守区域,设计了针对家畜衣原体的引物和探针,优化反应条件,建立家畜衣原体荧光定量PCR检测方法.结果显示:该方法的最低检测限为1.68×101 copies/μL,在1.68×102～1.68×106 copies/μL范围内具有良好的线性关系;该方法可特异性检测家畜衣原体,与肺炎衣原体、沙眼衣原体、鹦鹉热衣原体、流产衣原体4种4个菌株无交叉反应,表明特异性好;其批间重复试验和批内重复试验的变异系数分别为0.14％～0.51％、1.09％～1.49％,表明重复性良好.本方法可用于家畜衣原体的早期检测和流行病学调查.

Alfalfa mosaic virus（AMV） is a plant virus with worldwide distribution, wide host range, and serious harmfulness, which causes serious disease in soybean. In this study, the antiserum prepared by prokaryotic expression of AMV CP protein was used to establish an efficient and accurate indirect ELISA method for detection of AMV, which was applied to disease investigation and resistance identification. The results showed that the titers of the prepared three antiserums against recombinant protein and crude extract of soybean plants infected with AMV reached 256 000 times. Serum specificity analysis showed that the three antiserums only recognized AMV infected soybean leaves and did not recognize soybean mosaic virus（SMV） infected soybean leaves. A total of 50 soybean samples suspected to be infected with AMV were simultaneously detected by the established AMV indirect ELISA and conventional RT-PCR methods. The detection results of 46 samples were consistent, and the coincidence rate was 92%. The established AMV ELISA method and SMV ELISA method were used to detect the virus in soybean samples from the main soybean producing areas in Jilin province. The results showed that the detection rate of virus was 38.30%, the positive rate of SMV was 30.85%, the positive rate of AMV was 17.06%, and the composite infection rate was 9.61%. Resistance identification of 40 soybean varieties inoculated with AMV showed that all 40 soybean varieties were infected with AMV, but the viral load was significantly different. Some varieties showed AMV resistance, including 11 soybean resistant resources. The soybean disease caused by AMV had become one of the main diseases of soybean in Jilin province for the first time.

为了建立检测绵羊流产衣原体MOMP(main outer membrane protein)蛋白抗体的间接ELISA方法,试验将编码MOMP蛋白的基因序列经密码子优化后进行基因合成,并克隆和原核表达截短蛋白,以该蛋白为抗原建立绵羊流产衣原体抗体的间接ELISA方法,优化该方法的反应条件,同时验证其特异性、敏感性和重复性,最后用该方法对临床待检羊血清进行检测,并与间接血凝试验试剂盒的检测结果进行比较.结果表明:截短蛋白MOMP201～390的表达、纯化效果最好.经探索,优化后的间接ELISA方法最优反应条件:抗原包被浓度为1 mg/L,37℃包被2 h,待检血清最佳稀释度为1:100,血清和酶标二抗的最佳作用时间分别为45 min、60 min.建立的间接ELISA方法的敏感性较高,特异性和重复性较好,使用该方法从209份临床待检羊血清样本中检出47份阳性血清,与间接血凝试验试剂盒分别检测临床待检羊血清样本86份,总符合率为80.23％.说明基于截短蛋白MOMP201～390建立的绵羊流产衣原体抗体间接ELISA方法可用于绵羊流产衣原体临床血清抗体的检测及流行病学调查.

为了建立检测绵羊家畜衣原体血清抗体的间接ELISA方法,本试验构建pET-28b-ompA重组表达质粒,经诱导表达和纯化后,对重组主要外膜蛋白(MOMP)进行Western blot鉴定;以MOMP为包被抗原,通过反应条件筛选建立绵羊家畜衣原体血清抗体间接ELISA检测方法,分析其敏感性、特异性、重复性和符合性,并进行临床样品检测.结果显示,MOMP以包涵体形式表达,经Western blot鉴定重组蛋白具有良好的反应原性.优化反应条件为:抗原包被浓度为1 mg/L,37℃静置孵育1 h;血清抗体稀释倍数为1∶100,37℃反应1 h;酶标二抗稀释倍数为1∶5 000,37℃孵育1 h;底物显色条件为37℃避光5 min.利用52份绵羊血清确定ELISA方法的阳性判定临界值为P/N=2.156.该方法的敏感性、特异性和重复性较高.用该方法与间接血凝试验对86份血清进行检测,两者符合率为73.2％.用该方法对临床采集的205份绵羊血清样品进行检测,阳性率为34.1％.结果表明,本试验建立的绵羊家畜衣原体血清抗体间接ELISA检测方法具有较高的特异性和敏感性,可应用于临床检测.

In ovo immunization of chicken embryos with live vaccines is an effective strategy to protect chickens against various viral pathogens. The immunogenic efficacies of in ovo administration of lactic acid bacteria (LAB) in combination with live Newcastle disease (ND) vaccine were investigated in this study. Four hundred healthy 1-day-old fertilized specific pathogen-free (SPF) eggs of similar weights were randomly assigned to one of four treatments, with five replicates of each treatment and a total of 20 for each replicate. On day 18.5 of incubation, in ovo injections were given. The treatment groups are as follows: (I) no injection, (II) 0.9% physiological saline injection, (III) ND vaccine injection, and (IV) LAB as an adjuvant for ND vaccine injection. The ND vaccine adjuvanted with LAB significantly increased the daily weight gain, immune organ index, and small intestine histomorphological development in layer chicks while decreasing the feed conversion ratio (FCR). The results suggested that the LAB-adjuvant group significantly affected the relative expression of mucosal mucin protein (mucin-1) and zoccluding small circle protein-1 (ZO-1) (P < 0.05), whereas the relative expression of occludin mRNA was not significantly affected (P > 0.05) compared with the non-injected group. Meanwhile, we indicated that intra-amniotic synbiotic injection significantly maintained the balance of flora (P < 0.05). Compared with the non-injected group, the ND vaccine adjuvanted with the LAB group exhibited significant promotion of the HI and SIgA antibody titers in serum on day 21 (P < 0.05), induction of higher production of cytokines (IL-2, IL-4, IL-6, IFN-γ) in serum. In summary, in ovo injection of ND vaccine adjuvanted with LAB has a positive impact on the growth performance, immune function, and microbiome of growing chicks.

本研究利用小RNA深度测序法在大豆叶片上检测到1株花生斑驳病毒Peanut mottle virus (PeMoV),根据小RNA深度测序结果和GenBank公布的PeMoV基因组序列设计引物克隆了PeMoV公主岭分离物(PeMoV-Gongzhuling)的基因组序列.测序结果经拼接后获得了PeMoV-Gongzhuling基因组,大小为9709个核苷酸(Gen-Bank登录号:MT790744).该基因组在第123位—第9422位存在1个大的开放阅读框(ORF),编码1个多聚蛋白(分子量351.28 kD).PeMoV-Gongzhuling与GenBank中已登录的其他PeMoV分离物的基因组的核苷酸序列一致性为95.88％～99.53％,氨基酸序列一致性为97.39％～99.84％,其中与韩国分离物GYBU-92 (MT603819)在核苷酸和氨基酸水平上的一致性均为最高.

【Objective】In this study, monoclonal antibodies (MAbs) against the p30 protein of African swine fever virus (ASFV) were prepared and the linear epitopes on p30 was analyzed, which could lay the foundation for ASFV and its antibody detection as well as the study of p30 protein structure and function.【Method】BALB/c female mice aged 6 to 8 weeks were immunized with prokaryotic expression and purified recombinant p30 protein. The mice were immunized once every two weeks, with three times in total. First immunization was done with emulsification of antigen and equal volume Freund’s complete adjuvant, then, the mice were immunized with emulsification of antigen and equal volume of Freund’s incomplete adjuvant for the second and third immunization. After three immunizations, the tail was cut off and the blood was collected, and the serum antibody titer was detected by indirect enzyme-linked immunosorbent assay (ELISA). The mice with the highest serum titer were selected for enhanced immunization. Three days later, the mice spleen lymphocytes and SP2/0 myeloma cells were fused with PEG at a ratio of 4∶1. The positive hybridoma cells were screened by indirect ELISA by using the recombinant p30 protein as coated antigen. and the MAbs which could secrete antibodies steadily were cloned and purified by limited dilution method. ASFV was inoculated into porcine alveolar macrophages, and the indirect immunofluorescence assay (IFA) was performed with MAbs as primary antibody and rabbit anti-rat HRP-IgG as secondary antibody. The ASFV-infected and uninfected cells were precipitated and transferred to the nitrocellulose membrane by SDS-PAGE. p30 MAbs were obtained by Western blotting analysis of MAbs and identified positive by IFA, which was used as primary antibody and rabbit anti-rat HRP-IgG as secondary antibody. Primers were designed to amplify two truncated genes p30ab and p30bc, p30ab stands for truncated amino acid residues at position 86-153 and p30bc stands for truncated amino acid residues at position 120-153. The recombinant protein GST-p30ab and recombinant protein GST-p30bc were obtained by partial overlapping truncated p30 protein expression in prokaryotes. GST-p30ab and GST-p30bc fusion proteins were used as coated antigens, and the epitopes of p30 protein were preliminarily identified by indirect ELISA with 5 MAbs as primary antibodies, with rabbit anti-rat HRP-IgG as secondary antibodies.【Result】The purified recombinant protein was used as the coated antigen, and 25 hybridoma cell lines were screened by indirect ELISA, which could secrete anti-recombinant p30 protein. IFA results showed that 5 MAbs (8F4, 1D3, 1H2, 6C3 and 8E11) were positive for ASFV-infected cells. Western blotting results showed that all 5 strains of MAbs could react positively with ASFV-infected cells and negatively with uninfected cells. The recombinant p30 protein GST-p30ab was expressed in soluble and inclusion body forms, and GST-p30bc was expressed in inclusion body form. The truncated fusion proteins of the two groups were used as the coated antigens. Indirect ELISA showed that MAbs 8F4, 1H2 and 6C3 could effectively bind to the two recombinant proteins that means MAbs 8F4, 1H2 and 6C3 recognized the epitope amino acid 120-153; MAbs 8E11 and 1D3 could only bind to GST-p30ab protein, which meant they recognized the epitope amino acid 86-119.【Conclusion】In this study, the recombinant p30 protein with amino acid truncated at position 86-153 was expressed in soluble form. Five MAbs were prepared and two p30 protein epitopes were located. Combined with ELISA and IFA, it could be formed a very reliable method for the detection of ASFV and its antibody.

为获得非洲猪瘟病毒(African swine fever virus,ASFV)体外诱导表达的MGF505-3R蛋白.实验依据GenBank中报道的MGF505-3R基因序列设计引物,引入His标签,以ASFV基因全序列为模板扩增目的基因.将目的基因克隆至原核表达载体pGEX-6p-1,并转入大肠杆菌Transetta(DE3)中.用1 mmol/L异丙基 β-D-硫代半乳糖苷(IPTG)诱导表达菌体,分别取上清液和沉淀进行十二烷基硫酸钠-聚丙烯酰胺(SDS-PAGE)凝胶电泳分析.将表达的蛋白经过Ni-NTA琼脂糖亲和层析纯化.免疫印迹实验(Western-blot)分析蛋白的活性.克隆得到的MGF505-3R基因片段长度为843 bp,与原序列一致.蛋白在沉淀中以包涵体的形式表达,相对分子质量约为53 ku.纯化后可得到纯度较高的MGF505-3R重组蛋白.免疫印迹实验结果呈阳性.MGF505-3R蛋白可以在大肠杆菌Transetta(DE3)中表达并纯化,蛋白具有良好的表达活性.近几年,多基因家族(Multigene Families,MGFs)基因缺失疫苗被认为是目前ASF疫苗研制的主要方向,本研究获得的MGF505-3R重组蛋白将为ASFV MGF505-3R蛋白的研究提供基础,也为基因缺失疫苗相关免疫学方面的检测提供前期技术储备.

洋葱黄矮病毒(Onion yellow dwarf virus,OYDV)是危害葱属植物的主要病毒之一.本试验根据OYDV外壳蛋白(coat protein,CP)基因保守序列设计特异性引物,建立并优化了OYDV实时荧光RT-PCR检测方法,并对其特异性、敏感性和重复性进行了验证.结果显示:标准曲线Ct值与模板拷贝数的对数呈良好的线性关系,相关系数R2为0.996,扩增效率为95.921％;与青葱X病毒(Shallot virus X,SVX)、葱潜隐病毒(Shallot latent virus,SLV)均无交叉反应;最低检出限为2.0×102 copies· μL-1,比常规RT-PCR大约高1000倍;组内和组间变异系数均小于2％,具有良好的重复性.利用该方法对分别采集的各45份疑似OYDV感染的分蘖洋葱(珠葱)和大蒜样品进行检测,检出率较常规RT-PCR分别高40.00、6.67百分点,能更加客观地反映样品带毒情况.本试验建立的OYDV实时荧光RT-PCR方法具有良好的特异性、敏感性和重复性,可用于田间样品OYDV检测,为OYDV的有效防控提供技术支持.

为了获得非洲猪瘟病毒(African swine fever virus,ASFV)体外诱导表达的MGF360-13L蛋白,试验依据GenBank中报道的MGF360-13L基因序列设计引物,引入His标签,以ASFV全序列为模板扩增目的片段.将目的基因克隆到原核表达载体pGEX-6p-1上,转化BL21(DE3)大肠杆菌感受态细胞中进行体外诱导表达,聚丙烯酰胺凝胶电泳(SDS-P AGE)分析蛋白质的表达情况,将表达的重组蛋白通过镍琼脂糖凝胶柱亲和层析纯化,蛋白质印迹(Western-blot)分析蛋白质的活性.结果表明:克隆得到的MGF360-13L基因片段长度为1 060 bp,获得的重组蛋白在沉淀中以包涵体的形式表达,分子质量约为60 ku,纯化后可获得纯度较高的MGF360-13L,用BCA法测定最高浓度为2.73 mg/mL.Western-blot检测结果显示在60 ku处有一条明显的特异性条带说明MGF360-13L蛋白可以在BL21(DE3)大肠杆菌感受态细胞中表达并纯化,纯化后可获得纯度较高的MGF360-13L,且具有良好的反应原性.

马铃薯已成为我国第四大主粮作物,具有重要的战略意义,而马铃薯病毒病是主要的病害之一.本研究通过克隆马铃薯S病毒(potato virus S,PVS)的衣壳蛋白(Coat Protein)基因,连接表达载体pET28b,转化大肠杆菌,体外表达纯化出PVSCP.免疫日本大耳兔,制备出PVSCP多克隆抗体,其识别重组蛋白的效价为64 000倍,识别病毒的效价为32 000倍;抗体特异性分析表明,制备的多克隆抗体只识别PVS,不识别马铃薯M病毒、马铃薯Y病毒和马铃薯卷叶病毒.PVS多克隆抗体的制备,为马铃薯病毒检测方法提供了技术支持,为种薯质量的提高奠定了基础.

[目的]马铃薯Y病毒(potato virus Y,PVY)是影响马铃薯产量和品质的主要病毒之一,目前尚未发现有效的防治药剂,脱毒种薯的应用是预防PVY危害的主要防治措施.建立灵敏度高、特异性强的PVY快速检测方法,为脱毒种薯质量控制提供技术支撑.[方法]将PVY衣壳蛋白(coat protein,CP)分段表达为有重叠部分的小段多肽,以其为抗原通过Western blot分析PVY-CP的抗原表位.以P/N值最大为标准,通过常规双抗夹心ELISA(DAS-ELISA)对识别不同抗原表位的单克隆抗体(monoclonal antibody,MAb)进行配对试验,筛选一组检测效果最优的配对单克隆抗体,并确认捕获抗体和检测抗体.通过方阵滴定法确定捕获抗体及检测抗体的最佳工作浓度,通过控制变量法确定检测抗体与抗原的最佳共同孵育时间.以不同浓度的PVY-CP蛋白为抗原,检验快速DAS-ELISA的灵敏度.以感染不同病毒的马铃薯样品为抗原,检测快速DAS-ELISA的特异性.同时通过快速DAS-ELISA与RT-PCR检测50份田间采集的疑似感染PVY的马铃薯样品,将二者结果相比较检验检测方法的符合率.运用建立的快速DAS-ELISA对不同株系PVY感染的马铃薯样品进行检测.[结果]利用PVY-CP的6条分段表达多肽筛选到一组能进行DAS-ELISA的配对单克隆抗体(9G6和3D3),并以这对单抗为基础建立了 PVY快速DAS-ELISA检测方法.以9G6为捕获抗体包被酶标板,检测抗体3D3经辣根过氧化物酶(horseradish peroxidase,HRP)标记后以2 μg·mL-1的工作浓度与抗原在37℃共同孵育5 min.该检测方法检测限为0.5 ng·mL1,特异性分析结果显示该法仅在检测感染PVY的马铃薯样品时呈阳性反应,检测马铃薯S病毒(potato virus S,PVS)、马铃薯M病毒(potato virus M,PVM)、马铃薯卷叶病毒(potato leaf-roll virus,PLRV)等其他常见马铃薯病毒样品均呈阴性反应.通过快速DAS-ELISA与RT-PCR同时对50份田间采集的马铃薯样品进行检测,有48份样品检测结果一致,符合率达96％,且对PVY1及PVY0样品检测结果均呈阳性.[结论]建立的PVY检测方法灵敏度高、特异性强,30min即可完成检测,方便快捷,为PVY的高通量检测、脱毒种薯的生产提供了关键技术支持.

在吉林省7个主要甘薯种植区共采集85份甘薯叶片样品,利用小RNA深度测序技术对混合样品进行检测,经RT-PCR和测序验证,鉴定出样品中存在10种病毒,包括6种RNA病毒和4种DNA病毒.分别是马铃薯Y病毒科马铃薯Y病毒属的甘薯羽状斑驳病毒Sweet potato feathery mottle virus (SPFMV)、甘薯潜隐病毒Sweet po-tato latent virus (SPLV)、甘薯G病毒Sweet potato virus G(SPVG)、甘薯C病毒Sweet potato virus C(SPVC)、甘薯2号病毒Sweet potato virus 2(SPV2);长线形病毒科毛形病毒属的甘薯褪绿矮化病毒Sweet potato chlorotic stunt virus (SPCSV);双生病毒科菜豆金色花叶病毒属的甘薯曲叶病毒Sweet potato leaf curl virus(SPLCV);玉米线条病毒属的甘薯无症状1号病毒Sweet potato symptomless virus 1(SPSMV1);花椰菜花叶病毒科杆状DNA病毒属的甘薯杆状DNA病毒B Sweet potato badnavirus B(SPBV-B)和甘薯隐症病毒Sweet potato pakakuy virus(SPPV).

本研究克隆马铃薯卷叶病毒(Potato leafroll virus,PLRV)衣壳蛋白(Coat Protein,CP)基因,改造其密码子,使其偏好原核表达,构建pCzn1-PLRV CP重组表达载体,通过大肠杆菌表达纯化,经Western Blot方法鉴定为PLRV CP蛋白,纯化后的蛋白免疫日本大耳兔,成功制备PLRV CP蛋白多克隆抗体,识别重组蛋白效价为128000倍,识别PLRV叶片的效价为32000倍,经Western Blot方法分析,其特异性良好.本研究为PLRV检测方法的建立及脱毒种薯的检测提供了必需的生物材料.

随着转基因大豆(Glycine max)市场的扩大,我国对转基因大豆的食品安全、环境风险等问题争议日渐加剧.越来越多的国家要求对转基因食品进行标识,部分国家对转基因成分含量亦有要求.为了建立转Bar基因大豆的快速、有效检测方法,对转Bar基因大豆进行定量检测,本研究以纯化的膦丝菌素乙酰转移酶(phosphinothricin acetyltransferase,PAT)蛋白为抗原免疫小鼠(Mus musculus),成功制备了18株特异性良好的PAT单克隆抗体(monoclonal antibody,MAb),利用在大肠杆菌(Escherichia coli)中表达的PAT分段蛋白筛选到一组能进行夹心ELISA的配对单克隆抗体9F5和3G12,并以这对单抗为基础建立了PAT快速定量双抗夹心ELISA(double antibody sandwich ELISA,DAS-ELISA)检测方法.该方法以9F5为捕获抗体包被酶标板,检测抗体3G12经辣根过氧化物酶(horseradish peroxidase,HRP)标记后以2μg/mL的工作浓度与抗原在37℃共同孵育30 min,室温避光显色15～20 min.该检测方法检测限为1.69 ng/mL,对PAT蛋白的检测范围为0.5～8.0μg/mL,板内板间变异系数均小于10％.该法灵敏度高、稳定性好,1 h即可完成检测,为转Bar基因大豆的快速定量检测提供了技术支撑.

为了研究肉羊场羊只感染肠道寄生虫的病因及诊治方法,分别采用饱和蔗糖溶液漂浮法、饱和盐水漂浮法和碘染法对该养羊场39份粪便样本进行处理后镜检,并采用PCR方法对芽囊原虫进行检测、测序与分型.结果显示,球虫卵囊、阿米巴、毛首线虫卵、圆线虫卵、芽囊原虫阳性率分别为64.10％、12.82％、23.08％、5.13％、7.69％.两种以上寄生虫感染率为30.77％.6月龄以上羊的肠道寄生虫感染率显著大于6月龄以下羊.调查结果表明,该肉羊繁育基地肠道寄生虫感染普遍,应加强综合防疫,并对人兽共患基因型寄生虫的防治工作给予高度重视.

本研究为明确吉林和黑龙江省毛葱病毒病发生率,从两省5个地区共采集255份毛葱样品.根据毛葱4种主要病毒基因组序列设计特异性引物,对胡葱黄条病毒Shallot yellow stripe virus(SYSV)和青葱X病毒Shallot virus X(SVX)、洋葱黄矮病毒Onion yellow dwarf virus (OYDV)和葱潜隐病毒Shallot latent virus (SLV)进行双重RT-PCR检测.结果 表明,229份样品检出病毒,带毒率为89.8％,SLV的检出率最高,达87.06％,OYDV次之,为36.86％,SYSV检出率偏低,为0.78％;同时存在病毒复合侵染,其中双病毒复合侵染为SLV和OYDV,检出率为33.3％;三病毒复合侵染为SYSV、OYDV和SLV,检出率为0.78％,未发现4种病毒复合侵染.本研究为吉林和黑龙江种植区毛葱病毒病防治提供了参考依据.

中国每年进口大量转基因大豆(Glycine max),其中主要为cp4 epsps转基因大豆,需建立一种针对cp4 epsps转基因大豆的高效的快速ELISA鉴定方法.本研究利用合成多肽法将CP4-EPSPS蛋白分段表达,初步定位了5株CP4-EPSPS单克隆抗体识别的抗原表位,采用方阵法对识别不同抗原表位的单抗进行配对,以P/N最大值确定工作抗体及其浓度,通过控制变量法,确定最佳检测条件,建立cp4 epsps转基因大豆快速双抗夹心ELISA(double antibody sandwich ELISA,DAS-ELISA)检测方法.并通过特异性试验、重复性试验、阳性判定值试验,对建立的检测方法进行性能评估,利用建立的检测方法和商品化试剂盒同时对130份样品进行检测,比较其符合率.通过方阵法确定单抗2D3为捕获抗体,单抗1D10为检测抗体时检测信号最强,2D3工作浓度为20μg/mL,1D10工作浓度为10μg/mL.捕获抗体最佳包被条件为37℃封闭2 h,4℃过夜,样品与检测抗体先后加入酶标板37℃共同孵育10 min.该方法灵敏度为叶片或籽粒稀释160倍(g/mL),待检叶片最佳稀释范围是10～80(g/mL)、籽粒最佳稀释范围是10～40(g/mL).板内、板间变异系数小于25％,阳性判定值为1.41.对70份大豆叶片、40份大豆籽粒和20份豆浆进行检测,结果与CP4-EPSPS蛋白ELISA试剂盒检测结果比较,符合率为100％,且与其他蛋白不发生交叉反应.本研究建立的快速ELISA检测方法具有良好的准确性、重复性和特异性,该检测方法仅需30 min完成检测,适用于大豆植株、大豆籽粒及豆浆的快速鉴定.

为了快速检测转Cp4 epsps基因大豆,本研究以纯化后的CP4-EPSPS单克隆抗体1D12为捕获抗体,辣根过氧化物酶标记的羊抗CP4-EPSPS多克隆抗体8092为检测抗体,通过优化抗体工作浓度和抗原抗体反应时间,成功建立转Cp4 epsps基因大豆快速双抗夹心ELISA检测方法.结果显示:最佳检测条件为捕获抗体浓度10 μg/mL,检测抗体浓度1.25 μg/mL,样品与检测抗体先后加入酶标板37℃共同孵育60 min.该方法的检测范围为0.312 5～80 μg/mL,待检叶片、籽粒最佳检测范围为10～80倍稀释;板内变异系数为1.64％～5.42％,板间变异系数为3.05％～9.13％,符合ELISA定性试剂盒参考标准;对100份大豆叶片、20份大豆籽粒进行检测,与Western blot结果和标准结果进行比较,符合率为100％,表明该检测方法具有良好的准确性和重复性.该检测方法75 min内即可完成检测,适用于快速检测转Cp4 epsps基因大豆,为转Cp4 epsps基因快速检测试剂盒的开发奠定了基础.

为筛选防治玉米蚜虫的有效药剂,采用室内毒力测定与田间药效试验相结合的方法,研究了双丙环虫酯等复配剂对玉米蚜的防效.结果表明:双丙环虫酯与吡蚜酮质量比1:5时,共毒系数极值为172.20,有效成分用量为(10+50)g·hm-2对玉米蚜田间防效达86.59％;双丙环虫酯与吡虫啉质量比1:1时,共毒系数极值为183.80,有效成分用量为(15+15)g·hm-2对玉米蚜的田间防效达93.74％.丙环虫酯与噻虫啉质量比1:5时,共毒系数极值为196.12,有效成分用量为(10+50)g·hm-2对玉米蚜的田间防效达89.19％.双丙环虫酯与吡蚜酮、吡虫啉、噻虫啉复配防治玉米蚜具有增效作用,可有效地控制该虫的发生.