共查询到17条相似文献，搜索用时 109 毫秒
1.
[目的]构建适用于转基因大豆MON89788检测的质粒标准分子。[方法]利用定性PCR和连接、转化等分子克隆技术,将大豆内标准基因lectin、MON89788的3′端特异性序列和5′端特异性序列依次克隆到pMD18-T载体上,获得质粒标准分子pMD-LM3M5,并进行适用性验证。[结果]获得了3700bp的质粒标准分子,其中重组DNA片段1029bp。该质粒标准分子的定性PCR检测灵敏度达到10copy。[结论]该研究构建的质粒标准分子pMD-LM3M5能替代MON89788基体标准品,用于MON89788大豆及其产品的定性PCR检测。 相似文献
2.
转基因大豆MON89788转化体特异性定性PCR检测 总被引：6，自引：2，他引：4
[目的]建立MON89788大豆转化体特异性定性PCR检测方法。[方法]利用TAIL-PCR技术分离MON89788大豆的3′端旁侧序列,据此序列设计特异性引物进行PCR检测,并对该方法的特异性和灵敏度进行测试。[结果]获得了1142bp的3′端旁侧序列;依据该序列建立的PCR检测方法能特异性从MON89788大豆中扩增出170bp的产物,检测灵敏度达到0.05%,约为40个起始模板拷贝。[结论]该研究建立的方法特异性强、灵敏度高,可适用于MON89788大豆检测。 相似文献
3.
转基因大豆MON89788转化体特异性定性PCR检测（摘要）（英文） 总被引：1，自引：0，他引：1
[目的]建立MON89788大豆转化体特异性定性PCR检测方法。[方法]利用TAIL-PCR技术分离MON89788大豆的3′端旁侧序列,据此序列设计3条特异性引物用于旁侧序列的扩增。经过3轮PCR扩增,获得特异性扩增产物,将此产物回收后克隆到pMD-18T载体上测序,所得序列用Vector NTI分析,并提交NCBI进行序列比对。并对该方法的特异性和灵敏度进行测试。[结果]获得了1 142 bp的3′端旁侧序列。经Blast检索比对,该序列包括2部分,1 ～618 bp为载体序列（E9终止子部分序列和LB序列）,619 ～1 142 bp为大豆基因组序列;依据该序列建立的PCR检测方法能特异性地从MON89788大豆中扩增出170 bp的产物,检测灵敏度达到0.05%,约为40个起始模板拷贝。[结论]该研究建立的方法特异性强、灵敏度高,可适用于MON89788大豆检测。 相似文献
4.
金黄色葡萄球菌FnBP配体结合区基因的克隆及其原核表达(英文) 总被引：1，自引：0，他引：1
[Objective] The study aimed to clone the FnBP ligand binding gene of Staphylococcus aureus and run prokaryotic expression by constructing a prokaryotic expression vector. [Method] The gene encoding FnBP ligand binding gene was amplified from S.aureus chromosomal DNA by PCR technique. After T-A cloning, plasmid pMD18- FnBP was constructed. pMD18- FnBP and pET28a(+)were digested by BamH Ⅰ and EcoR Ⅰ double enzymes, then the purified FnBP ligand binding gene was subcloned into the expression vector pET28a(+), and the prokaryotic expression vector pET28a-FnBP was thus constructed. The constructed plasmid pET28a-FnBP was transformed into Escherichia coli BL21(DE3) competent cells. The bacterium was induced by IPTG and the expressed products were analyzed by SDS-PAGE and Western blot. [Result] The gene fragment with the length of 370 bp was amplified by PCR approach. One approximately 30 kD exogenous protein was observed in SDS-PAGE analysis. Western blot analysis indicates the protein has antigenicity of S.aureus. [Conclusion] The FnBP ligand binding gene of S.aureus was successfully cloned and expressed in prokaryotic cells. 相似文献
5.
[目的]研究牛瑟氏泰勒虫P23表面蛋白基因的克隆及原核表达.[方法]采用PCR方法扩增牛瑟氏泰勒虫中国延边株P23基因片段,将扩增产物克隆人pMD18-T载体构建重组质粒pMD18-P23经PCR、双酶切鉴定后测序;将目的基因片段亚克隆人表达载体pGEX-4T-1构建重组表达质粒pGEX-4T-P23,转化宿主菌BL21获得重组菌.通过对诱导条件的优化,根据SDS-PAGE确定表达蛋白的最佳表达条件;Western-blotting检测表达蛋白的反应原性.[结果]所克隆的牛瑟氏泰勒虫P23基因片段长507 bp,与牛瑟氏泰勒虫日本株P23基因的核苷酸同源性达99.4%,表达的融合蛋白大小约为46 ku;诱导时机以接种培养后2 h为最佳,诱导时间以6 h为最佳,诱导温度以34℃为最佳,0.008～1.000 mmol/L的IPTC对表达量的影响不大.Western blotting检测表明该蛋白具有较好的抗原性.[结论]为牛瑟氏泰勒虫病的免疫学诊断和预防等研究奠定了基础.
Abstract：
[Objective] The aim was to study cloning and prokaryotic expression of P23 major surface protein gene of Theileria sergenti.[Method]A pair of specific primers was designed according to the sequence of P23 major surface protein of T.sergenti (D84447).The P23 gene was amplified by PCR from genomic DNA of T.sergenti and cloned into pMD18-T vector to construct recombinant clonal vector pMD18-P23.Positive clones were identified by PCR screening and restriction digestion.A recombinant expression plasmid pGEX-4T-P23 was constructed by subcloning the cloned P23 gene into the linearized pGEX-4T-1 vector and transformed into E.coil BL21.After introduction by IPTG,the expressed fusion protein was identified by SDS-PAGE and Western-blotting. [Result] The cloned gene has a total length of 507 bp.Sequencing result showed that the nucleotide sequence of the cloned P23 gene shared 99.4% identity with that of P23 published in GenBank (D64447).The expressed fusion protein was 46 ku in molecular mass.Induction opportunity of zhours after culture inoculation was the best,the induction time of 6 h was the best,and induction temperature of 34 ℃ was the best as well,IPTG of 1 mmol/L had little effect on the expression.Western-blotting indicated that recombinant protein was recognized by specific antibody. [Conclusion] This study would lay a foundation for further research on the prevention and diagnose of T,sergenti. 相似文献
6.
TaqMan quantitative PCR technique was used to detect the copies of exogenous CaMV35S flanks sequence in transgenic soybean. With soybean lectin as the endogenous reference gene, and gene complex DNA in... 相似文献
9.
双价抗虫转基因大豆抗苜蓿夜蛾分析(英文) 总被引：4，自引：1，他引：3
[Objective] The aim of the research was to analyze the resistance of binary insect-resistant transgenic soybean to Heliothis viriplaca.[Method]In this experiment, resistance analysis of the stabilized binary insect-resistant transgenic soybean to Heliothis viriplaca was conducted in lab and in field conditions.[Result] The results indicated that the leaves of insect-resistant transgenic soybeans T5-150 and T5-195 showed lighter damage than those of non-transgenic soybeans. Meanwhile, the Heliothis viriplaca larvae fed on leaves of these two transgenic soybeans were characterized by less leaf consumption, shortening survival day, slower development and less pupation.[Conclusion]It was concluded that insect-resistance of transgenic soybean to Heliothis viriplaca was increased dramatically and the research provided a reference for selecting binary insect-resistant transgenic soybean to Heliothis viriplaca. 相似文献
10.
Chen Xin Yan Jiyong Gao Bing Peerasak srinives 《农业科学与技术》2006,7(1):8-13
A study was conducted in the field of the Institute of Vegetable Crops, Jiangsu province from July 2000 to August 2003. The transgenic roundup-ready soybean was sown in the middle of the field in a circular manner for 5 circles, with the distance of 3 m, from one circle to another. Then the wild soybean was planted in plots as the rays of the circles in 8 directions （N, E, W, S, NE, NW, SE and SW）, spaced every 5 m until 50 m. Each plot comprised 25 plants. In the second year, the wild soybean seeds from the first year were planted in the field together with the original wild soybean as check. Before flowering time, high concentrations of roundups （about 4-5 times of the normal dose） were sprayed on the plants and the surviving plants were identified. The leaves were taken to the lab for DNA extraction to determine the unique DNA for roundup-ready soybean （CTAB method）. About 2% of the plants survived, but some leaves were yellow. One plant of wild soybean was found to have the roundup-ready gene from the original roundup-ready soybean. The other surviving wild soybeans should also had some fragments of the roundup tolerance gene. However, the DNA bands were not very clear in the PCR map. 相似文献
11.
牛体外发育胚胎特定阶段差异表达基因的研究 总被引：1，自引：0，他引：1
[目的]研究不同发育时期牛体外受精胚胎在基因表达模式上的差异.[方法]利用单个胚胎mRNA差异显示技术,对单个8细胞期胚胎与囊胚进行mRNA差异显示,获得1条特异表达条带,对其进行克隆、测序,并与GenBank进行对比.[结果]该序列与牛核糖体蛋白131基因(ribosomal protein 131,RPL3])具有99%的同源性.采用实时定量PCR技术检测8细胞期和囊胚期胚胎RPL31的mRNA表达量,结果表明,RPL31在8细胞期胚胎的相对表达量为囊胚期胚胎的3.2倍.[结论]为揭示和阐明控制牛早期胚胎发育的相关机理提供依据.
Abstract：
The cattle different stage embryos obtained from in vitro was studied using the technology of single preimplantation embryo mRNA different display:single 8-cell and blastocyst stage embryos were studied using technology of mRNA different display and one different fragment was found.The result suggested that this fragment displayed high homology (99%) to cattle mRNA for ribosomal protein L31.Then to detect the expression of RPL31 mRNA in 8 cell and blastocyst stage embryos by real-time quantitative PCR,the result showed the relative amount of 8 cells was 3.2 times of blastocyst's. 相似文献
12.
[目的]构建抗黄瓜花叶病毒RNAi载体,并将载体转入烟草.[方法]采用RT-PCR方法,扩增黄瓜花叶病毒NS04加工番茄分离物的RNA2基因组的序列选取CMV RAN2基因组中的复制酶片段作为靶序列,构建pBi35SCR2真核表达载体,并对表达载体时行鉴定;通过农杆菌介导的方法将表达载体转入烟草,用PCR的方法检测载体是否转入.[结果]系统进化树分析结果表明,RNA2中编码CMV-2a的序列与中国浙江的DQ412731 分离物有较高核苷本乡酸及氨基酸同源性,分别达到98.0%和96.5%;RCR结果表明,试验成功构建了pBi35SCR2真核表达载体,并成功将表达载体转入烟草[结论]试验获得的转基因烟草可作为后期攻毒试验的材料,并为研究加工番茄抗黄瓜花叶病毒奠定了基础.
Abstract：
[Objective] Aimed to construct RNAi vector resistant to cucumber mosaic virus and transferred this vector into tobacco.[Method]RT-PCR method was used to amplify cucumber mosaic virus NSO4 and process RNA2 gene sequen of tomato isolates.The analysis results of phylogenetic tree demonstrated that the sequence in RNA2 encoded CMV-2a had 98.0% and 96.5% homology with nucleotide and amino acid of DQ412731 isolate of Zhejiang,China.The replicase fragment in CMV RAN2 gene was taken as target sequence to construct pBi35SCR2 eukaryotic expression vector,then the expression vector was identified.Through agrobacterium-mediated method,the expression vector was transferred into tabacco and PCR method was used to check the transfer.The PCR results demonstrated that the experiment had successfully construct eukaryotic expression vector of pBi35SCR2 and the expression vector was successfully transferred into tabacco. [Conclusion] The obtained transgenic tobacco could be used as challenge test material in following experiment and provided foundation for studying processing tomato resist cucumber mosaic virus. 相似文献
13.
底栖鱼类对水田上覆水中磷素动态的扰动效应 总被引：2，自引：0，他引：2
[目的]研究底栖鱼类泥鳅对水田上覆水中磷素动态的扰动效应,探讨生物扰动机制.[方法]基于模拟试验,使用离子色谱法和分光光度法,对比分析上覆水中磷素含量在有/无泥鳅活动时的差异.[结果]扰动组的TP、DTP和PP浓度在试验开始阶段与对照组无显著差异,在试验中、后期显著高于对照(P<0.05).扰动组要的PP/TP高于对照组,扰动组中TP浓度的增加主要是由于PP的增加,扰动组的DIP/DT在试验中、后期显著高于对照(P<0.05).[结论]底栖鱼类对水田上覆水中的磷素产生了扰动作用,增加了水稻生长可利用的的磷素养分.
Abstract：
[Objective] The research aimed to investigate the bioturbation effects of benthic fish Misgurnus anguillicaudatus on phosphorus dynamic in overlying water of paddy field,as well as to explore the bioturbation mechanism.[Method]Based on simulation experiment,the phosphorus contents in overlying water were analyzed comparatively with and without Misgurnus anguillicaudatus by the using of ion chromatography and spectrophotometry. [Result] The concentrations of total phosphorus (TP),dissolved total phosphorus(DTP)and particular phosphorus(PP) in bioturbation group had no significant differences with those in control group in initial stage of experiment,and became significantly higher than control group in middle and late stages of experiment(P<0.05).The PP/TP ratios in bioturbation group were bigger than those in control group,the increase of TP concentration in bioturbation group was mainly due to the increase of PP.The ratios of dissolved inorganic phosphorus(DIP) to DTP (DIP/DTP) were significantly bigger than those in control group in middle and late stages of experiment (P<0.05). [Conclusion] The benthic fish had bioturbation effects on phosphorus in overlying water of paddy field,which increased the available phosphorus for rice growth. 相似文献
14.
一株产紫杉醇的曼地亚红豆杉内生真菌的分离及鉴定 总被引：1，自引：0，他引：1
[目的]分离并鉴定1株产紫杉醇的曼地亚红豆杉内的内生真菌.[方法]从曼地亚红豆杉树皮内表皮中分离得到32株内生真菌,并通过高效液相色谱法检测其发酵产物.[结果]筛选获得1株可以产紫杉醇的内生真菌M57,其紫杉醇产量为45～50μg/L,并通过对M57菌落的形态学观察以及18S rDNA序列分析初步将其鉴定为根霉属(Rhizopus)真菌.[结论]该菌株的发现为微生物发酵法生产紫杉醇提供了具有潜在应用价值新的菌种.
Abstract：
[Objective] The aim was to isolate and identify a taxol-producing endophytic fungus from Taxus media.[Method]32 strains of endophytic fungi were identified form the inner bark of T.media,and their fermentation products were detected by high performance liquid chromatography (HPLC). [Result] Through the screening,a strain of taxol-producing endophytic fungi M57 was obtained,which could produce 45-50 μg/L of taxol,and M57 was defined as Rhizopus sp.through morphological observation and 18S rDNA sequence analysis. [Conclusion] The finding of Rhizopus sp.M57 provided a promising strain for producing taxol with taxol-producing fungi fermentation process. 相似文献
15.
依据最新NDB数据库中蛋白质-DNA复合物晶体结构数据,基于修正的DNA结构统计力学模型,利用蒙特卡洛多重积分计算DNA动力学结构的有关参数,并对计算得到的结果进行时间复杂度和精确度分析.
Abstract：
Based on protein-DNA complex crystal structural data in up-to-date Nucleic Acid Database,the related parameters of DNA Kinetic Structure were investigated by Monte-Carlo Multiple Integrals on the base of modified DNA structure statistical mechanical model,and time complexity and precision were analyzed on the calculated results. 相似文献
16.
干旱胁迫对苗期甘蔗叶片水分和叶绿素荧光参数的影响 总被引：6，自引：0，他引：6
[目的]研究干旱胁迫对甘蔗叶片水分和叶绿素荧光参数的影响,为甘蔗生产及评价研究提供依据.[方法]选取7个抗旱性不同的甘蔗品种,在苗期进行干旱胁迫,并测定胁迫条件下甘蔗叶片水分含量和叶绿素荧光参数的变化.[结果]甘蔗叶片水势和相对含水量与土壤相对含水量存在一定的内在联系,耐旱强的品种对土壤水分的利用率较高;相关分析和因子分析表明茁期干旱存活率、Fc/Fm、叶片水势和相对含水量可被用作抗旱性评价指标.[结论]水势表现为一个相对独立的影响因子,对甘蔗抗早性有支配作用,并验证了Fv/Fm作为甘蔗抗旱评价指标的可靠性.
Abstract：
[Objective] The aim was to explore the effects of water stress on leaf water and chlorophyll fluorescence parameters of sugarcane seedling,as well as to provide basis for the study on sugarcane production and evaluation.[Method]Seven different sugarcane varieties were studied at the seedling stage under drought stress,and the changes of leaf water and chlorophyll fluorescence parameters under stress conditions were detected. [Result] leaf water potential,leaf relative water content and soil relative water content showed a certain amount of internal relationship,the sugarcane varieties that had more tolerant to drought had higher utilization rate of soil water;the correlation analysis and factor analysis suggested that the survival rate at seedling stage under drought stress,Fv/Fm,leaf water potential and relative water content could be used as drought resistance evaluation indicators. [Conclusion] As a relatively independent influencing factor,water potential had dominating effect on drought resistance,and the reliability of Fv/Fm as drought resistance evaluation indicator had been verified. 相似文献
17.
中草药提取液对黄瓜苗期杀根结线虫的活性研究 总被引：1，自引：0，他引：1
[目的]研究中草药提取液对黄瓜苗期杀根结线虫的活性.[方法]利用植物水培技术,结合生长条件易控制、根系生长过程易连续观察的特点,应用5种具有较强室内触杀根结线虫活性的植物提取液及其与阿维菌素的复配液,对黄瓜苗期水培根系杀线活性进行系统研究.[结果]胡黄连和石榴皮提取液可以在不影响水培黄瓜苗期植株生长的前提下表现出对根结线虫良好的活体植株根系寄生防治和杀灭活性,且达到了与阿维菌素相近的作用水平;而狗脊、木香和蛇床子提取液对黄瓜的生长表现出不同程度的抑制作用,其对根结线虫的防治效用和杀灭活性也较弱.[结论]触杀效果和杀线活性成分能否被植物吸收利用共同制约着具有杀线活性中草药提取液的开发应用.
Abstract：
Present mature plants hydroponic technology was used,combined with some excellent characteristics,such as growth conditions was easy to control and process of root growth was easy to continuously observe,the nematicidal activity of 5 kinds of Chinese herbs extracts and the compound solution of Avermectin,with strong contact toxicity effect indoor,was systematically studied and investigated the affection on the rootknot nematode parasitized on the cucumber seeding stage.It is found that under the premise of no influence on root growth of cucumber,extracts from Picrorhiza scrophulariiflora and Punica granatum showed strong prevention and nematicidal activity,and had the similar efficacy of Avermectin;while the extracts from Cibotium barornetz,Aucklandia lappa Decne and Fructus cnidii showed low nematicidal activity and various degrees inhibition effect on plant growth. 相似文献