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1.
为了进一步了解CHI基因在青稞黄酮合成中所起的作用,以青稞品系94-19-1为材料,通过同源克隆技术分离CHI基因,并对分离得到的CHI基因进行生物信息学分析及原核表达。结果表明,从青稞94-19-1中克隆得到的CHI基因编码区长为696bp,编码231个氨基酸。序列分析表明,青稞CHI基因有4个碱基和大麦不同,导致1个氨基酸发生改变。SDS-PAGE检测结果表明,将该基因克隆到表达载体pET-32a上,并转化大肠杆菌BL21后,经诱导可成功表达出融合蛋白。 相似文献
2.
为进一步研究查尔酮合酶（Chalcone synthase,CHS）在黄酮化合物代谢过程中的作用,以青稞94-19-1为材料,通过同源克隆技术分离CHS基因,并对分离得到的CHS基因进行生物信息学分析及原核表达。结果表明,从青稞94-19-1中克隆得到的CHS基因编码区长为1 197 bp,编码398个氨基酸。生物信息学分析表明,该基因编码的蛋白质分子量为43.479 kDa,预测等电点(pI)为5.92,是酸性蛋白;该酶蛋白体外红细胞中的半衰期为30 h,不稳定系数(II)大于40,属于不稳定蛋白;平均亲水指数为-0.090,是亲水性的蛋白。SDS-PAGE检测结果表明,将该基因克隆到表达载体pET-32a上,并在大肠杆菌BL21中表达,可得到64 kDa左右的融合蛋白,在IPTG诱导浓度为1.0 mmol·L-1时,最佳诱导时间为3 h。 相似文献
3.
获得了一类茶树氧甲基转移酶基因cDNA全长并构建了该基因的原核表达载体。以从茶树叶片中提取的总RNA为模板,结合RT-PCR与RACE克隆技术获得氧甲基转移酶(O-methyltransferase)基因cDNA全长1 280 bp,其开放阅读框为1 068 bp,编码355个氨基酸,推测的蛋白分子量为39.1 kD,理论等电点为5.68。其氨基酸序列与葡萄和蓖麻氧甲基转移酶基因相似性分别为73%、71%。将该基因片段连接到原核表达载体pET-28a中,转化大肠杆菌BL21后诱导重组蛋白的表达,经IPTG诱导,SDS-PAGE检测到1条与预测融合蛋白分子量相符的外源蛋白。 相似文献
4.
茶树α-tubulin基因的原核表达及其多克隆抗体的制备 总被引：1，自引：0，他引：1
根据茶树α-微管蛋白(α-tubulin)的mRNA全长序列(GenBank登录号DQ340766)设计引物,以RT-PCR方法扩增茶树α-tubulin基因,将扩增产物克隆到原核表达载体pET-32a(+)上,并转化至大肠杆菌BL21trxB(DE3)中,经异丙基-β-D硫代半乳糖苷(IPTG)诱导后以包涵体形式表达。以纯化后的α-tubulin融合蛋白制备兔多克隆抗体,用Westernblot方法检测抗体特异性。结果显示α-tubulin蛋白以包涵体形式大量表达,以融合蛋白制备的抗体对茶树体内的α-tubulin具有良好的特异性。 相似文献
5.
为了给转HARDY(HRD)基因小麦研究奠定基础,利用PCR技术从拟南芥中克隆HRD基因,进行生物信息学分析,预测其编码的蛋白质结构与功能,构建原核表达载体pET28a(+)-HRD,转化大肠杆菌BL21(DE3),IPTG诱导蛋白的表达,同时构建pBin-HRD植物表达载体。测序分析结果表明,克隆的HRD与NCBI发布的拟南芥核苷酸序列的同源性为99%,其开放阅读框长555bp,可编码184个氨基酸,具有许多重要功能位点;成功构建了原核表达载体pET28a(+)-HRD和植物表达载体pBin-HRD,诱导表达出大小约为23.3kD的蛋白,与理论值相近。 相似文献
6.
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
7.
黄酮类化合物在植物中参与过滤紫外线、固氮和花色形成等过程,异黄酮对人体有抗氧化、预防乳腺癌等保健作用。查尔酮还原酶(chalcone reductase,CHR)是植物中参与黄酮类化合物代谢的重要酶。克隆大豆查尔酮还原酶基因并构建植物表达载体,有助于进一步研究其功能和异黄酮的代谢过程。采用RT-PCR方法,从栽培大豆(Gly-cine max)南农1138-2中,克隆得到了第14号染色体上的一个编码大豆查尔酮还原酶(chalcone reductase,CHR)的基因,命名为GmCHR。该基因含有948 bp长的编码区序列(Coding DNA Sequence,CDS),编码315个氨基酸。预测其蛋白质分子量为35.5 kDa,等电点为6.32。与其他豆科植物中的查尔酮还原酶相比,GmCHR蛋白序列与葛藤(Puerari-ae montana)CHR的相似性最高,达94%。组织表达分析表明,在自然生长条件下GmCHR在叶中的表达量最大;其次是种子;在花和茎中相同;在根中的表达量最小。利用Gateway方法获得植物过表达载体pMDC83-GmCHR,经检测表明过表达载体已成功转化农杆菌EHA105,为今后进一步了解GmCHR在大豆异黄酮代谢过程中的功能提供材料基础。 相似文献
8.
水稻瘤矮病毒S9基因的生物信息学分析及原核表达蛋白的抗血清制备 总被引：1，自引：0，他引：1
以广东德庆市水稻瘤矮病的典型病株为材料,RT-PCR扩增并克隆了水稻瘤矮病毒(Rice gall dwarf virus,RGDV)的第9组分全长核苷酸序列。生物信息学分析表明,其基因结构特征与已报道的其他分离物基本一致。Pns9蛋白为细胞膜外蛋白,在植物呼肠孤病毒属内没有发现同源蛋白,与霍乱弧菌(Vibrio cholerae)的入口蛋白(portal protein)及姜氏军团菌(Legionella drancourtii LLAP12)的假设蛋白(hypothetical protein LDG_3259)分别有33%和24%的氨基酸序列相似性,功能尚待确定。将S9基因克隆至原核表达载体pET-28b(+)得到高效表达,融合蛋白经亲和层析纯化后,免疫新西兰大白兔制备抗血清。Western blot和ELISA分析表明制备的兔抗为特异抗血清,效价为1∶13 1072。 相似文献
9.
水稻黑条矮缩病病毒外壳蛋白基因S10的原核表达、多克隆抗体制备及应用 总被引：5，自引：2，他引：5
用RT-PCR方法从感染水稻黑条矮缩病毒(rice black-streaked dwarf virus,RBSDV)水稻中克隆该病毒的外壳蛋白基因S10,然后将此外壳蛋白基因再亚克隆到原核表达载体PET-32a中构建成重组原核表达载体pET32a-CP。将重组表达载体转化大肠杆菌BL21(DE3),经IPTG诱导,Ni+ NTA亲和柱纯化获得分子量约为76kD含硫氧还蛋白的融合蛋白。以纯化的重组蛋白为抗原免疫兔子制备RBSDV外壳蛋白的多克隆抗体,并用制备的多克隆抗体建立了可靠、灵敏、特异的检测RBSDV的免疫捕获RT-PCR及Dot-blot ELISA方法,为该水稻病毒病的诊断提供技术支持。 相似文献
10.
茶树无色花色素还原酶基因克隆及表达分析 总被引：7，自引：3，他引：4
采用EST测序技术和3′RACE技术,获得了茶树儿茶素代谢途径中的一个重要酶—无色花色素还原酶的全长基因,在GenBank的登录号为EF205148,序列全长1 301 bp,其中开放阅读框长1 029 bp,编码342个氨基酸,3′端有一个明显的多聚腺苷酸加尾信号,推测的蛋白分子量约为37.5 kD,理论等电点为5.81。将该基因重组到表达载体pET-32a(+)中进行原核表达,经IPTG诱导、SDS-PAGE检测,结果表明茶树无色花色素还原酶基因能在大肠杆菌BL21中表达,电泳检测到一条大约60 kD的外源蛋白,与预测的融合蛋白分子量相符。同源性分析表明茶树无色花色素还原酶基因编码的氨基酸序列与其他植物具有较高的相似性,例如与亚洲棉、草莓和葡萄的相似性分别为70%、68%、71%。利用半定量PCR技术检测总儿茶素含量不同的4个茶树品种中与类黄酮合成相关的黄酮醇合成酶(FLS)、二氢黄酮醇-4-还原酶(DFR)、无色花色素还原酶(LAR)、花色素合成酶(ANS)等7个基因的表达情况,结果表明DFR和LAR基因的表达量与茶树中总儿茶素含量呈一定的相关性,而其他基因则与其相关性不大。 相似文献
11.
12.
《Journal of Crop Improvement》2013,27(2):81-114
Summary The efficiency of N fertilizers is usually poor; often less than 50% of the applied N is taken up by the crop. This review focuses on various N fertilizers with respect to the significance of different N loss pathways, namely (i) ammonia volatilization, (ii) dinitrogen and nitrogen oxide emissions, and (iii) nitrate leaching. Further, the significance of biological N immobilization, ammonium fixation and, finally, the impact of nitrate vs. ammonium uptake on crop yield are also discussed. The reviewed literature shows that N fertilizers may differ markedly in their susceptibility to losses. There is, however, considerable scope to improve N efficiency of each N source by proper N management practices. 相似文献
13.
Lakshman Lal 《Potato Research》1987,30(2):329-334
Summary The effects of the leaves of five plant species, one from each of the generaAmbrosia, Anemone, Eupatorium, Eucalyptus andLantana, on potato tuber moth were investigated under indigenous storage conditions at the Central Potato Research Station, Shillong
(1800 m above sea level). Their action was compared with that of a biological insecticide (spores ofBacillus thuringiensis), a chemical insecticide (carbaryl), and an untreated control. The data collected after six months storage on tuber damage,
sprout damage and the rotting indicated that the leaves ofLantana aculeata provided most protection to the tubers, reducing damage from over 70% in the check to below 5%, and sprout damage from over
45% to below 3%. Next best wasEucalyptus globulus followed byB. thuringiensis. They may be used on tubers stored for table use or for seed as they had no adverse effect on germination or on the yield
of a subsequent crop. 相似文献
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15.
Joseph E. Laferriere Charles W. Weber Edwin A. Kohlhepp 《Plant foods for human nutrition (Dordrecht, Netherlands)》1991,41(3):277-282
Teas of plant origin traditionally consumed by the Mountain Pima of Chihuahua, Mexico, were analyzed for mineral nutritional content. Fe, Cu, Zn, Ca, and Mg composition was determined for native teas made from shoots ofTagetes lucida, T. filifolia, Elytraria imbricata, andHolodiscus dumosus, and from root xylem ofCeanothus depressus andPhaseolus ritensis. Native uses of these teas are also described. 相似文献
16.
Novel food and non-food uses for sorghum and millets 总被引：4，自引：3，他引：4
Sorghum and millets have considerable potential in foods and beverages. As they are gluten-free they are suitable for coeliacs. Sorghum is also a potentially important source of nutraceuticals such antioxidant phenolics and cholesterol-lowering waxes. Cakes, cookies, pasta, a parboiled rice-like product and snack foods have been successfully produced from sorghum and, in some cases, millets. Wheat-free sorghum or millet bread remains the main challenge. Additives such as native and pre-gelatinised starches, hydrocolloids, fat, egg and rye pentosans improve bread quality. However, specific volumes are lower than those for wheat bread or gluten-free breads based on pure starches, and in many cases, breads tend to stale faster. Lager and stout beers with sorghum are brewed commercially. Sorghum's high-starch gelatinisation temperature and low beta-amylase activity remain problems with regard to complete substitution of barley malt with sorghum malt . The role of the sorghum endosperm matrix protein and cell wall components in limiting extract is a research focus. Brewing with millets is still at an experimental stage. Sorghum could be important for bioethanol and other bio-industrial products. Bioethanol research has focused on improving the economics of the process through cultivar selection, method development for low-quality grain and pre-processing to recover valuable by-products. Potential by-products such as the kafirin prolamin proteins and the pericarp wax have potential as bioplastic films and coatings for foods, primarily due to their hydrophobicity. 相似文献
17.
M. I. Siri P. Villanueva M. J. Pianzzola L. Franco Fraguas G. Galván M. Acosta F. Ferreira 《Potato Research》2004,47(3-4):127-138
Summary The in vitro antimicrobial activity of extracts from accessions ofSolanum commersonii Dun. collected in the south of Uruguay was investigated against five microorganisms including the pathogenRalstonia solanacearum. A total of 30 extracts corresponding to organic and aqueous extracts were studied. Interestingly, most of the positive results
for growth inhibition were againstR. solanacearum. The extracts were also analyzed for the presence of glycoalkaloids and lectins. Six of the ten aqueous extracts showed lectin
presence and a wide variation in the type and amounts of glycoalkaloids, was found. Results indicate that there is no clear
relationship between the antimicrobial activity against the five microorganisms screened and the presence or amounts of lectins
and glycoalkaloids, traditionally regarded as possible antimicrobial metabolites in theSolanum genus, which indicates the presence of as yet unidentified antimicrobial compounds. 相似文献
18.
Alejandro Carlos Tozzini María Fernanda Ceriani María Verónica Saladrigas H. Esteban Hopp 《Potato Research》1991,34(3):317-324
Summary Clones derived from thirty-one different accessions (nineteen of Argentine origin) belonging to eightSolanum species were screened for resistance to infection by potato virus X strain cp (PVX cp) by mechanical inoculation of plantlets
that had been micropropagated in vitro. Estimates of PVX multiplication obtained by enzyme linked immunosorbent assay and
slot blot nucleic acid hybridization allowed the identification of resistant clones derived from five accessions belonging
toS. commersonii S. oplocense, S. sparsipilum andS. tuberosum andigena. Resistant genotypes supported PVX concentrations 5 to 15 times smaller than did the susceptible control cultivar Spunta.
Graft inoculation test confirmed the presence of extreme resistance similar to that conferred by the ‘immunity’ gene X1 (also called RXact). 相似文献
19.
Rita Favas Janaína Morone Rosrio Martins Vitor Vasconcelos Graciliana Lopes 《Marine drugs》2022,20(3)
The loss of density and elasticity, the appearance of wrinkles and hyperpigmentation are among the first noticeable signs of skin aging. Beyond UV radiation and oxidative stress, matrix metalloproteinases (MMPs) assume a preponderant role in the process, since their deregulation results in the degradation of most extracellular matrix components. In this survey, four cyanobacteria strains were explored for their capacity to produce secondary metabolites with biotechnological potential for use in anti-aging formulations. Leptolyngbya boryana LEGE 15486 and Cephalothrix lacustris LEGE 15493 from freshwater ecosystems, and Leptolyngbya cf. ectocarpi LEGE 11479 and Nodosilinea nodulosa LEGE 06104 from marine habitats were sequentially extracted with acetone and water, and extracts were analyzed for their toxicity in cell lines with key roles in the skin context (HaCAT, 3T3L1, and hCMEC). The non-toxic extracts were chemically characterized in terms of proteins, carotenoids, phenols, and chlorophyll a, and their anti-aging potential was explored through their ability to scavenge the physiological free radical superoxide anion radical (O2•−), to reduce the activity of the MMPs elastase and hyaluronidase, to inhibit tyrosinase and thus avoid melanin production, and to block UV-B radiation (sun protection factor, SPF). Leptolyngbya species stood out for anti-aging purposes: L. boryana LEGE 15486 presented a remarkable SPF of 19 (at 200 µg/mL), being among the best species regarding O2•− scavenging, (IC50 = 99.50 µg/mL) and also being able to inhibit tyrosinase (IC25 = 784 µg/mL), proving to be promising against UV-induced skin-aging; L. ectocarpi LEGE 11479 was more efficient in inhibiting MMPs (hyaluronidase, IC50 = 863 µg/mL; elastase, IC50 = 391 µg/mL), thus being the choice to retard dermal density loss. Principal component analysis (PCA) of the data allowed the grouping of extracts into three groups, according to their chemical composition; the correlation of carotenoids and chlorophyll a with MMPs activity (p < 0.01), O2•− scavenging with phenolic compounds (p < 0.01), and phycocyanin and allophycocyanin with SPF, pointing to these compounds in particular as responsible for UV-B blockage. This original survey explores, for the first time, the biotechnological potential of these cyanobacteria strains in the field of skin aging, demonstrating the promising, innovative, and multifactorial nature of these microorganisms. 相似文献
20.
Alejandro M. S. Mayer Abimael D. Rodríguez Orazio Taglialatela-Scafati Nobuhiro Fusetani 《Marine drugs》2013,11(7):2510-2573
The peer-reviewed marine pharmacology literature from 2009 to 2011 is presented in this review, following the format used in the 1998–2008 reviews of this series. The pharmacology of structurally-characterized compounds isolated from marine animals, algae, fungi and bacteria is discussed in a comprehensive manner. Antibacterial, antifungal, antiprotozoal, antituberculosis, and antiviral pharmacological activities were reported for 102 marine natural products. Additionally, 60 marine compounds were observed to affect the immune and nervous system as well as possess antidiabetic and anti-inflammatory effects. Finally, 68 marine metabolites were shown to interact with a variety of receptors and molecular targets, and thus will probably contribute to multiple pharmacological classes upon further mechanism of action studies. Marine pharmacology during 2009–2011 remained a global enterprise, with researchers from 35 countries, and the United States, contributing to the preclinical pharmacology of 262 marine compounds which are part of the preclinical pharmaceutical pipeline. Continued pharmacological research with marine natural products will contribute to enhance the marine pharmaceutical clinical pipeline, which in 2013 consisted of 17 marine natural products, analogs or derivatives targeting a limited number of disease categories. 相似文献