共查询到20条相似文献，搜索用时 656 毫秒
1.
以连接在pMD-18T载体上的香蕉条斑病毒云南分离物(BSV)全基因组为模板,设计特异引物,PCR扩增获得ORFⅠ全长序列。目的片段及原核表达载体pET32(b)分别经EcoRⅠ和XhoⅠ双酶切后,连接构建重组质粒pET32-ORFⅠ,将鉴定为正确的重组质粒转化E.coli BL21(DE3)感受态细胞,获得原核表达OFRⅠ工程菌。工程菌在28℃条件下,用浓度为0.1 mmol/L的IPTG诱导表达6 h,离心收集菌液并超声波破碎,获得以含目的肽段的可溶性蛋白为主的融合蛋白。将融合蛋白通过Ni2+-NTA亲和层析柱纯化后,得到纯度较高的融合蛋白。以此蛋白为抗原免疫家兔,获得特异抗血清,该抗血清的效价达1∶100 000,Western Blot验证表明,抗血清的特异性较强。本研究为BSV的快速检测以及BSV编码蛋白的功能研究奠定了基础。 相似文献
2.
香蕉束顶病毒(Banana bunchy top virus,BBTV)是危害香蕉的主要病毒之一,严重威胁着香蕉的生产.BBTV基因组至少由6个环状ssDNA组分组成,其中DNA2组分变异最大,是否编码蛋白及其功能缺乏研究.本研究通过PCR方法克隆了BBTV Haikou2 DNA2ORF,将其构建到pET32a载体中,进行原核表达和抗血清制备.结果表明重组表达载体pET32a-DNA2ORF在表达菌Transetta(DE3)中表达一个约21 ku的特异融合蛋白,通过超声波破碎仪破碎细胞和诱导条件优化,分析表明,该蛋白主要以可溶形式存在,最佳表达条件为30℃、1.0 mmol/L IPTG诱导3h.融合蛋白经过Ni2+-NTA琼脂糖亲和层析纯化,免疫家兔制备出抗血清.Western blot实验表明抗血清具有特异性; 酶联法(ID-ELISA)测定表明,抗血清效价达到25 000,能够检测出香蕉汁液中加入的0.954 μg/mL浓度的表达纯化蛋白. 相似文献
3.
小麦热激蛋白WHSP90基因的原核表达及多克隆抗体制备 总被引：1，自引：0，他引：1
为了进一步研究HSP90基因的功能,将WHSP90基因构建到原核表达载体pET-28a(+)中,得到His-WHSP90融合表达载体,并转化到大肠杆菌BL21(DE3)中,发现1 mmol/L IPTG诱导4 h蛋白表达量最大;经过蛋白标记亲和层析柱(HisTrapTMHP)纯化得到的纯化蛋白浓度达到0.4 mg/mL.免疫家兔制备抗体,用间接ELISA法检测免疫后家兔抗血清效价大于125 000,满足后续试验要求的效价值,为在蛋白水平上研究WHSP90基因功能提供了基础. 相似文献
4.
《中国糖料》2014,(2)
根据已发表ScYLV-P0基因系列设计特异性引物,应用RT-PCR技术从甘蔗病叶的mRNA扩增得到目的 DNA片段。以pET32a(+)为原核表达载体,构建重组表达质粒pET32a-P0。经过双酶切鉴定和DNA测序后,将重组表达质粒转入大肠杆菌BL21(DE3)pLySs,在30℃培养条件下IPTG诱导表达。通过SDS-PAGE电泳检测融合蛋白表达情况。表达结果显示,在该表达系统中,融合表达蛋白P0是以包涵体形式的蛋白存在;P0融合蛋白大小约45kDa,与P0开放阅读框的理论推算值29.991 kDa加上载体自身蛋白约18 kDa相符,用Ni2+-NTA琼脂糖亲和层析纯化融合蛋白,免疫家兔制备出抗血清,通过酶联法(ID-ELISA)测定本实验制备的ScYLY-P0抗血清工作浓度为1∶25000。 相似文献
5.
应用PCR方法从感染香蕉束顶病毒的香蕉植株幼嫩假茎和叶片总DNA中克隆了Rb(Rb-binding-like protein)基因的编码区,将其插入原核表达载体pET-28b中构建重组质粒pET28b-Rb。转化重组质粒的E.coli BL21(DE3)进行IPTG诱导表达,SDS-PAGE分析结果表明,表达产物大小为22.6ku,且主要以包涵体形式稳定表达。目的蛋白经Ni2+-NTA琼脂糖亲和层析纯化后免疫家兔并获得抗血清。Western blot检测结果表明,抗血清与诱导表达的BBTV编码Rb蛋白发生特异性反应。间接ELISA法测定的抗血清效价大于1:250000。用抗血清对感病香蕉和健康香蕉进行检测,结果表明,抗血清对感病香蕉有很高特异性,最佳工作浓度为1:1500。 相似文献
6.
类黄酮7-O-甲基转移酶(F7-OMT)具有甲基转移酶功能,能催化底物类黄酮甲基化产生植物抗毒素,提高植物的抗菌性。为了给进一步研究该酶蛋白的功能奠定基础,以青稞"94-19-1"为材料,采用同源克隆方法得到青稞F7-OMT基因编码片段。生物信息学分析表明,青稞F7-OMT开放阅读框(ORF)全长为1173bp,编码390个氨基酸,蛋白分子质量为42 207.8Da,等电点pI为5.36,无信号肽序列,具有甲基转移酶2(Methyltransf_2,pfam00891)和二聚化(Dimerization,pfam08100)保守域。通过亚克隆将F7-OMT连接到表达载体pET-32a上,构建pET-32a-F7-OMT融合表达载体,在E.coli BL21(DE3)pLysS中诱导表达,并采用亲和层析纯化法得到纯化蛋白。 相似文献
7.
8.
通过PCR扩增的方法从大豆品种Harosoy中克隆到1个新的b ZIP基因(Gmb ZIP71),基因表达分析表明Gmb ZIP71在叶片、生长点、花、花芽、荚和根等多个器官中表达。将Gmb ZIP71构建到原核表达载体p ET29b上,导入大肠杆菌BL21(DE3)中,对其进行IPTG诱导。结果表明:在IPTG浓度为0.25 mmol·L-1,诱导时间为4 h,诱导温度为28℃时,重组蛋白得到表达,分子量大约为48 k Da,SDS-PAGE电泳结果表明重组蛋白主要以包涵体形式存在,用His蛋白纯化系统回收得到Gmb ZIP71-His重组蛋白,EMSA(electrophoretic mobility shift assay)实验表明Gmb ZIP71重组蛋白可以在体外与ACGT顺式作用元件结合。 相似文献
9.
水稻黑条矮缩病毒基因组S7编码的2个非结构蛋白在病株中的表达检测 总被引：1，自引：1，他引：0
水稻黑条矮缩病毒（RBSDV）基因组片段S7含有2个非重叠的阅读框,所编码的蛋白分别为p7a和p7b。根据RBSDV S7序列（EU111804）设计特异性引物分别扩增编码p7a和p7b蛋白的基因片段,并亚克隆至原核表达载体pET 32a（+）和pSBET上,再以大肠杆菌BL21(DE3)pLysS或BL21(DE3)pLysE为宿主菌进行高水平表达,利用纯化的蛋白免疫小鼠,制备了p7a和p7b蛋白的特异性抗血清。蛋白质印迹分析表明p7a和p7b蛋白均在水稻病株中表达,但p7a含量较为丰富,易于在病株中检测到,而p7b在病株中含量则很低;在病毒粒子中,两者均未检测到任何信号,表明两者均为RBSDV的非结构蛋白。 相似文献
10.
对大豆东农42喷施表油菜素内酯(epi BL),进行GmRAV定量,研究油菜素内酯(BR)对GmRAV基因表达的影响,同时对T5代GmRAVox烟草外源添加epi BL,研究GmRAV基因是否也参与BR信号从而抑制植物的生长发育。结果表明:BR抑制GmRAV基因表达,并且外源添加epiBL并未促进GmRAVox烟草植株茎延伸,表明GmRAV可能为BR促进植物的生长发育信号传导途径的负调节因子。同时,将大豆中已克隆的GmRAV基因克隆到原核表达载体p ET28a上,构建与His标签融合的重组蛋白pET28a-GmRAV质粒,转化大肠杆菌BL21(DE3)中,对其进行37℃、28℃两种不同温度下0.5mmol·L~(-1)IPTG诱导,诱导时间为6 h。SDS-PAGE电泳结果表明:分子量大约为40 kDa的重组蛋白在不同温度下均表达,低温可以明显增强GmRAV蛋白的可溶性。 相似文献
11.
12.
2017—2018年,在同安镇粮食产能区示范推广水稻病虫害绿色防控与统防统治融合技术模式,取得了显著成效。融合技术示范区比常规区减少用药2~3次,用药量减少37.6%,防治成本降低400元/hm^2;水稻产量增加12.8%,收益增加2 835元/hm^2。 相似文献
13.
《Journal of Crop Improvement》2013,27(1):313-324
Summary Arbuscular mycorrhizae were inoculated into phosphorus-deficient soil fertilized with either organic or chemical fertilizer with cucumber (Cucumis sativus L.) as the first crop and lettuce (Lactu-ca sativa L.) as the second crop but without additional fertilization and AM inoculation. AM increased dry matter and fruit yield of cucumber significantly in the unfertilized, organic-fertilized and P-deficient plants compared with the fully chemical-fertilized plants. AM inoculation increased the available phosphorus in plant and soil by around 30% for all treatments except for those chemically-fertilized. The rate of AM infection did not differ significantly among the fertilization treatments, but the infection intensity was higher in unfertilized, organic-fertilized and phosphorus-deficient treatments than chemical-fertilized treatment. The residual effects of AM-inoculated to cucumber were evident for lettuce in all pre-treatments that were unfertilized and un-inoculated for the second cropping. Without P-fertilization, neither crop could grow optimally even when the soil was inoculated with AM, suggesting that AM could not serve as a substitute for phosphorus fertilizer. However, the other beneficial effects of AM on crop growth and yield could not be fulfilled with phosphorus fertilizer. 相似文献
14.
为明确春季调控措施对冬小麦生长发育和籽粒产量的影响,以冬小麦品种石新828为材料进行田间试验,4个处理分别为:起身期追全部氮肥(除基肥外,下同)并叶面喷多效唑(N1);起身期追2/3氮肥并喷多效唑+拔节期追1/3氮肥(N2);起身期追1/3氮肥+拔节期追2/3氮肥(N3);拔节期追全部氮肥(N4)。生育期间测定群体和个体生育特性,成熟期调查产量性状。结果表明,N1和N2处理小麦拔节期的总茎数、叶面积指数(LAI)和干物质积累量均显著高于N4和N3处理。孕穗期N4和N3处理小麦的总茎数、LAI和干物质积累量显著高于N1和N2处理。开花到成熟期各处理的总茎数、LAI和干物质积累量差异均不显著。孕穗期前,不同处理的株高差异不显著,孕穗期后,N4处理的株高最高,且显著高于N1处理。各处理基部节间直径和中上部节间长的差异不显著,N4处理基部第一节间长度显著大于其他处理。随追氮时期前移或前期施氮量增多,不孕小穗数减少,结实小穗和穗粒数增加,N1比N4处理不孕小穗数显著减少,结实小穗和穗粒数显著增多。N1处理小麦成熟期的千粒重最高,且显著高于N4处理。N1处理的籽粒产量最高,且显著高于N3和N4处理。起身期追氮配合多效唑调控,可以获得比拔节期追氮更高的穗粒数和千粒重,从而获得更高的产量。 相似文献
15.
百香果是海南省的新兴产业,也是我国多个省区扶贫攻坚的重要产业。病虫害是影响百香果产业的重要因素,随着海南百香果种植面积越来越大,病虫害对百香果产业的不利影响越来越凸显。由于缺乏百香果病虫害种类调查鉴定的基础性研究,许多种植户对田间发生的病虫害种类识别不足,从而影响了百香果病虫害的防控效果。为了摸清海南百香果种植区的病虫害种类,于2019—2021年对海南省儋州市、乐东县、屯昌县、文昌市、三亚市、东方市、保亭县、白沙县、海口市等百香果种植区的病虫害进行了系统调查与鉴定,并对其发生危害特点进行了研究。鉴定发现海南百香果种植区的病害有11种,分别是病毒病、茎基腐病、炭疽病、疮痂病、疫病、灰霉病、果腐病、褐斑病、烟煤病、藻斑病和缺镁。重要病害为病毒病和茎基腐病,危害程度分别为4级和3级,炭疽病和疮痂病危害也较重,危害程度均为2级,其他病害危害较轻。害虫有28种,分别是黄胸蓟马、茶黄蓟马、红带蓟马、美洲棘蓟马、普通大蓟马、咖啡木蠹蛾、美洲斑潜蝇、橘小实蝇、斜纹夜蛾、双线盗毒蛾、绿丽刺蛾、白带锯蛱蝶、烟粉虱、温室白粉虱、柑橘粉蚧、木槿曼粉蚧、藤壶蜡蚧、茶角盲蝽、樟颈曼盲蝽、稻绿蝽、中稻缘蝽、茶小绿叶蝉、丽球瓢蜡蝉、绿鳞象甲、眼斑芫菁、白条豆芫菁、甘薯梳龟甲和朱砂叶螨。重要害虫为黄胸蓟马和咖啡木蠹蛾,危害程度均为3级,茶黄蓟马和美洲斑潜蝇的危害也较重,危害程度均为2级,其他害虫危害较轻。软体动物3种,分别是灰巴蜗牛、同型巴蜗牛和一种尚未鉴定的蛞蝓。本研究明确了海南百香果种植区病虫害具体种类,新发现百香果病害1种和害虫24种,为海南百香果病虫害的精准防控提供技术支撑。 相似文献
16.
17.
Textile fibers were obtained from secondary polyethylene terephtalate (PET) and its mixtures with primary PET at initial orientation of 18000–33000 %, rate of additional orientation drawing 3.5–6.5 times and temperature of thermal fixation 363–413 K. The fibers’ tensile strength was found to decrease and elongation at break to increase with the decrease of their linear density under the conditions of fibers formation. For the fibers based on polymer mixtures, the presence of oxidized fragments in the secondary PET limited the compatibility of the two polymers which resulted in deteriorated tensile properties. The linear density (4–16 dtex), tensile strength (30–50 cN/tex) and elongation at break (20–60 %) of the PET fibers obtained were close to these for the industrially produced polymer fibers. The values of the average diameter of the fibers formed and oriented under laboratory conditions allows classifying them between the fine and the coarse textile fibers which makes them suitable for the textile industry. 相似文献
18.
《Plant Production Science》2013,16(4):281-285
AbstractTwo cultivars of spinach and one cultivar of komatsuna were sand cultured using a nutrient solution with Na and K in various combinations. The growth (fresh weight of shoot) of spinach cv. Atlas was significantly increased when 1.4 mM KC1 of the 7 m M KO was replaced with 1.4 mM NaCl, although the growth was reduced when more KC1 was replaced with NaCl. In another spinach cv. All Right, however, NaCl substituting for 20% of KC1 showed no significant effects. In komatsuna, Na used in place of K at any ratio reduced the growth. The growth of spinach cv. Atlas was increased by applying 20 mM NaCl, irrespective of K dose (480 - 2400 mg K2O per 1/2000 a Wagner pot). The growth of spinach cv. All Right was also increased by 20 mM NaCl when K was at a deficient level (480 - 960 mg K20 per pot), but was not when K was at: an excessive level (1920 - 2400 mg K2O per pot). In komatsuna, 20 mM NaCl reduced the growth. In spinach, Na prevented the decrease of the photosynthetic rate and stomatal conductance caused by K deficiency. 相似文献
19.
Gas-exchange characteristics of potato (Solanum tuberosum L.) canopies were determined with crop enclosures under condition of optimal water supply, during soil water depletion, and during recovery from drought-stress. The plants were grown under a rainshelter in large containers with 2-m2 surface area. Control plants received water throughout the growth period. In Expt. 1, transient drought was imposed by interrupting water supply completely for 22 days. During the 33-day period of drought treatment in Expt. 2, the stressed plants received half the amount of water that was given to the control plants (irrigations 2–3 times per week).
During soil water depletion, stomatal conductance of the canopy was reduced earlier and more than photosynthesis. This resulted in a lower internal carbon dioxide concentration in the leaves of stressed plants and a higher ratio between photosynthesis and stomatal conductance. The value of that ratio increased as soil water potential dropped.
Water-use efficiency ( ) increased during drought stress. The cultivar Bintje showed a larger ratio between photosynthesis and stomatal conductance of the canopy and a higher than cv. Saturna, both for optimal water supply and during drought stress.
Photosynthesis and conductance of the canopy started to recover immediately upon rewatering, even when relative values had declined to 20–30% during the preceding stress period. After release from stress, the leaves in the top layer of the canopy had a higher stomatal conductance in treated plants than in control plants. 相似文献
20.
本文建立了气质联用仪测定稻田环境中苯醚甲环唑和嘧菌酯的残留分析方法,对苯醚甲环唑和嘧菌酯在水稻、土壤和田水中的消解动态和残留规律进行了研究。苯醚甲环唑和嘧菌酯在糙米、稻壳、植株、土壤和田水中的最低检测浓度均为0.20 mg/kg,最小检出量为0.2 ng,在不同样品中的平均加标回收率为80.8%~109.5%,相对标准偏差在1.7%~19.7%之间。田间试验结果表明,苯醚甲环唑和嘧菌酯在水稻植株和田水中的残留消解动态规律均符合一级动力学反应模型,苯醚甲环唑在水稻植株和田水中的残留消解半衰期分别为6.3~11.6 d和1.4~11.6 d;嘧菌酯在水稻植株和田水中的残留消解半衰期分别为3.6~8.7 d和2.9~23.1 d。以推荐剂量600 g/hm2和1.5倍推荐剂量900 g/hm2,最多施药3次,距最后一次施药15 d时,苯醚甲环唑和嘧菌酯在糙米中的最高残留量分别为0.461 mg/kg和0.634 mg/kg,低于我国国家标准《食品中农药最大残留限量》(GB2763-2014)中规定的糙米中苯醚甲环唑最大残留限量0.5 mg/kg和欧盟、美国规定的糙米中嘧菌酯最大残留限量5.0 mg/kg。 相似文献