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1.
欧李叶片全长cDNA文库的构建和部分克隆的序列分析 总被引：4，自引：1，他引：3
采用SMART技术,构建了欧李(Prunus humilis Bunge)品系T8-1叶片全长cDNA文库,原始文库滴度达6.17×10~7 pfu/mL,库容达3.7×10~7,重组率达95%,平均插入片段大小约为1.2 kb.随机挑取400个单克隆进行5'端测序,共获得364个欧李ESTs.绝大部分ESTs的长度在500～1300 bp之间,平均长度为877 bp,其中具有完整ORF结构的序列有221条,占60.71%.通过与NCBI等非冗余核酸数据库和蛋白质数据库进行比对、查询和注释,获得已知功能基因或具推测功能的基因181个(339个ESTs),相似性较低的未鉴定基因5个(9个ESTs),新基因14个(16个ESTs). 相似文献
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扇贝性腺和肠组织表达序列标签的初步识别* 总被引：2，自引：0，他引：2
构建了扇贝(Placopecten magellanicus)的性腺和肠组织cDNA文库,并对96个克隆进行部分测序,获得82个表达序列标签(ESTs)序列.序列同源性搜索结果表明,72个EST序列与GenBank中的已知序列具有很高的同源性(e<10-8).其中19个序列属于rRNA基因;53个序列分别属于44个不同的基因.在44个基因中,有36个是编码已知功能的蛋白质基因,8个与未知功能的开放阅读框具同源性(假定蛋白);此外,44个基因全部是首次在扇贝中鉴定的基因;基因的功能分类显示,36个已知功能的基因分别属于基因表达与蛋白质合成、细胞结构、物质运输、代谢、细胞调节和信号传导等6大类. 相似文献
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利用甜菜夜蛾(Spodoptera exigua)围食膜多克隆抗体免疫筛选草螟(Loxostege sticticalis)中肠cDNA表达文库,得到编码肉碱-脂酰肉碱转位酶(LstiSLC25)基因的全长cDNA克隆.该cDNA克隆全长1 474 bp(GenBank登录号EU924506),开放阅读框长897 bp,编码299个氨基酸,预测分子量和等电点分别为32.1 kD和9.46.序列含有3个典型的溶质运转重复结构,具有溶质运载蛋白家族的典型特征.将该基因与pET30载体重组后,经IPTG诱导,蛋白在大肠杆菌(Escherichia coli获得了表达. 相似文献
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为挖掘花生茎中重要的功能基因,了解花生茎生长发育的分子机理,以闽花6号为材料,利用SMART技术成功构建了花生茎不同生育时期混合的全长cDNA文库,并对文库的部分序列进行了测序及分析。结果表明,该文库原始库容为1.25×106cfu·mL-1,重组率达100%,插入片段大小为750~2 000 bp,平均插入片段大小为1 000bp,达到文库质量要求。随机挑选50个单克隆进行5'端测序,共获得50条有效序列。通过与NCBI非冗余蛋白质数据库进行比对和注释,获得已知功能基因或具推测功能的基因29个,未知功能基因9个,新基因12个,其中47个(94%)基因为花生未报道的基因。因此,该文库的构建为克隆和研究花生茎部重要表达基因,从分子水平上揭示花生茎的生长发育规律提供了基础。 相似文献
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利用甜菜夜蛾Spodoptera exigua（Hübner）围食膜多克隆抗体免疫筛选草地螟（Loxostege sticticalis L．）中肠cDNA表达文库,得到编码运载蛋白LstiSLC25的全长cDNA克隆。该cDNA克隆全长1399bp(GenBank登录号EU924506),开放阅读框长897 bp,编码299个氨基酸,预测分子量和等电点分别为32.1kD和9.46。序列含有三个典型的溶质运转重复结构,具有溶质运载蛋白家族的典型特征。将该基因与pET30载体重组后,经IPTG诱导,蛋白在大肠杆菌中获得了表达。 相似文献
6.
甘薯多酚氧化酶cDNA的克隆及序列分析 总被引：2，自引：0，他引：2
植物多酚氧化酶（olyphenol oxidase,PPO)是一类核编码的铜金属酶，含有2个保守的铜结合区域（CuA和CuB)，N端和C端部分缺乏明显的同源性。根据植物PPO的结构特点，从2个保守的铜结合区域的氨基酸序列设计一对简并引物，扩增了甘薯（Ipomoea batatas L.) PPO保守区之间的cDNA片段，测序表明该片段含595bp，编码195个氨基酸，推导的氨基酸序列包含了植物PPO的2个保守的铜结合区域，与葡萄（Vitis vinifera)，番茄（Lycopersicon esculentum)马铃薯（Solanum tuberosum)和蚕豆（Vicia faba)PPO保守区的同源性分别为71．3％，80．8％，79．8％，70．6％。在此基础上设计引物通过3'RACE和5'RACE的方法获得了甘薯PPO cDNA的3'和5'端的片段。最后根据保守区的cDNA片段，3'RACD片段和5'RACE片段的序列信息进行拼接，推导出甘薯PPO cDNA的全部序列信息。结果表明：甘薯PPO cDNA全长含1984bp，有完整的阅读框，编码588个氨基酸。另外5'非翻译区16bp,3'非翻译区202bp，其中包括1个多聚腺苷酸化信号AATAAA，以及1个含17个腺苷酸的ploy(A)尾。由甘薯PPO cDNA推导的氨基酸邓列由葡萄，番茄，马铃薯和蚕豆PPO cDNA推导的氨基酸序列进行比较，它们之间存在较明显的同源性，同源性分别为49％，48％，47％和49％。 相似文献
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利用稻瘟病菌(Magnaporthegriesa)连续6个发育时期的材料构建了一个混合cDNA文库。文库滴度,重组率和插入片段长度等质量分析表明,构建的文库包含完整的稻瘟病菌基因,可用于病菌基因表达分析。利用该文库获得了7456条5′端表达序列标签(ESTs)(GenBank收录号:(CK909944￣CK913666和CK928583￣CK932582),生物信息分析表明:EST序列拼接出2975个假定独立转录本(TUTs),冗余度为60.1%;从cDNA文库中筛选出大量的低丰度表达基因,约占TUT总数的79.8%,说明在文库中基因组成类型的复杂性较高;在所有TUTs中,功能未知基因约占85.5%,编码ECM33蛋白和疏水蛋白等病菌致病相关的注释基因高丰度表达,进一步表明该cDNA文库反映了病菌侵染和发育过程中基因表达的状况。 相似文献
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松墨天牛（Monlchamus alternatus Hope）是林业重大害虫。本研究运用SMART技术构建了松墨天牛幼虫cDNA文库,库容量为1.09×10^6,文库滴度为5.45×10^6 pfu/mL,重组率为97.8%,平均插入片段大小约为1 000 bp,构建的文库质量较好。随机选取1 022个克隆进行测序,获得了1 012条表达序列标签（ESTs）,序列在GenBank中登录号为JZ143720～JZ144731。EST序列经过聚类拼接得到411条非重复单一序列,其中216条序列有功能注释。在216条有功能注释的序列中,有181条（83.80%）参与＂分子功能＂,122条（56.48%）参与＂细胞成分＂,168条参与＂分子生物学过程＂（77.78%）。研究结果为克隆和分析松墨天牛新的功能基因奠定了基础。 相似文献
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茶树β-葡萄糖苷酶cDNA克隆和原核表达 总被引：9，自引：0，他引：9
摘要：对与萜烯类香气前体及与抗病虫害有密切关系的茶树(Camellia sinensis ) β-葡萄糖苷酶cDNA进行克隆和原核表达。结果表明，该酶cDNA全长序列为1 475 bp(GenBank登录号为AF537127)，与其它植物同源性为40%～60%。α-螺旋构象14.33%、β-折叠构象25.43%，存在多个氨基酸功能结构域。利用pET-32a表达载体构建的重组质粒，转化到大肠杆菌(Escherichia coli )菌株BL21zztrxB（DE3）中，诱导产生63 kD的融合蛋白，表达产物具有正常的生物学活性，能催化葡萄糖苷键的水解反应；诱导表达结果显示，主要在细胞质中以可溶性蛋白形式进行表达。 相似文献
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海藻糖-6-磷酸合成酶(trehalose-6-phosphate synthase,TPS)是海藻糖合成途径中的一个关键酶.目前,TPS基因的研究多数集中于细菌和真菌等,而对植物的研究较少.本实验通过对茶树(Camellia sinensis(L.)O.Kuntze)全器官转录组文库序列比对,获得一条与其他物种同源性较高的编码TPS基因的EST序列,通过RACE扩增后获得茶树TPS基因cDNA全长序列,命名为Cs TPS(GenBank登录号JQ742017).该基因cDNA全长3 125 bp,包含一个2799 bp的开放阅读框,编码932个氨基酸.多序列比对分析结果表明,Cs TPS基因编码的蛋白具有明显的TPS和TPP两个结构域.系统进化分析表明,其编码的氨基酸序列与拟南芥(Arabidopsis thaliana)、烟草(Nicotiana tabacum)和番茄(Solanum lycopersicum)等植物的TPS同源性较高,且CsTPS与拟南芥TPS1(AtTPS1)的同源性高于TPS2(AtTPS2)和TPS3(AtTPS3).qPCR分析显示,CsTPS基因在茶树不同组织器官中呈现差异性表达.低温诱导促使老叶和嫩叶中的CsTPS基因上调程度明显大于根系,表明CsTPS基因可能参与了茶树抗寒机制. 相似文献
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Identification of oxidation products of (-)-epigallocatechin gallate and (-)-epigallocatechin with H(2)O(2) 总被引：4，自引：0，他引：4
Zhu N Huang TC Yu Y LaVoie EJ Yang CS Ho CT 《Journal of agricultural and food chemistry》2000,48(4):979-981
(-)-Epigallocatechin gallate (EGCG) and (-)-epigallocatechin (EGC) are two important antioxidants in tea. They also display some antitumor activities, and these activities are believed to be mainly due to their antioxidative effects. However, the specific mechanisms of antioxidant action of tea catechins remain unclear. In this study are isolated and identified two novel reaction products of EGCG and one product of EGC when they were reacted separately with H(2)O(2). These products are formed by the oxidation and decarboxylation of the A ring in the catechin molecule. This study provides unequivocal proof that the A ring of EGCG and EGC may also be an antioxidant site. This study also indicates an additional reaction pathway for the oxidation chemistry of tea catechins. 相似文献
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Roy A 《Journal of agricultural and food chemistry》1999,47(12):5209-5210
Terpinolene oxide, a monoterpene belonging to the p-menthane group, is easily derived from naturally abundant (R)-limonene. It was isomerized with montmorillonite clay catalyst to karahanaenone (2,2, 5-trimethylcyclohept-4-en-1-one) by ring enlargement. The enantiomers of the corresponding alcohol, karahanaenol (2,2, 5-trimethylcyclohept-4-en-1- ol), known for their individual organoleptic properties, were resolved through Pseudomonas cepacia lipase mediated enantiospecific alcoholysis of its acetate derivative. 相似文献
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W R Dantzer J Hopper K Mullin S Hendrich P A Murphy 《Journal of agricultural and food chemistry》1999,47(10):4291-4296
14C-Fumonisin B(1) (FB(1)) was produced by Fusarium proliferatum M-5991 in modified Myro liquid medium and purified to >95% purity with a specific activity of 1.7 mCi/mmol. Nine male and nine female F344/N rats were each dosed by gavage with 0.69 micromol of (14)C-FB(1), (14)C-hydrolyzed FB(1), or (14)C-FB(1)-fructose/kg body weight. Urinary excretion of (14)C-FB(1) and (14)C-FB(1)-fructose was 0.5% and 4.4% of the total dose, respectively, and was similar between male and female rats. Urinary excretion of (14)C-hydrolyzed HFB(1) was significantly greater (P > 0.05) in female rats as compared with male rats (17.3% vs 12.8% of the total dose, respectively). There were no significant (P > 0.05) differences in biliary excretion of the three fumonisin compounds with a mean of 1. 4% of the dose excreted at 4 h after dosing. Lesser amounts continued to be excreted up to 9.25 h after dosing. Although biliary excretion of the (14)C-FB(1), (14)C-hydrolyzed FB(1), and (14)C-FB(1)-fructose was similar, increased urinary excretion of the (14)C-hydrolyzed FB(1) as compared to (14)C-FB(1) and (14)C-FB(1)-fructose indicated a greater absorption of the hydrolyzed form. 相似文献
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S. R. Verma J. P. S. Arora J. S. Shankar R. Chand 《Water, air, and soil pollution》1987,36(3-4):247-257
The effect of protein oxovanadium(V) ion concentration and pH on the ratio of diffusion current (id/id0) was studied in vanadium(V) ovalbumin-S and denatured ovalbumin systems. In both the cases marked decrease in diffusion current was observed at the respective pH values, indicating that binding takes place with cationic groups of the proteins. The binding sites (n) were found to be pH dependent. The uniformity of logK and ΔG 0 value at all pH values indicated the involvement of same sites in interaction. Furthermore, the linear scatchard plots in both the systems supported the involvement of single class of independent sites in oxovanadium(V) anion interaction. The difference in binding sites (n) has been attributed to the folded structure of ovalbumin-S while unfolded one of denatured ovalbumin. 相似文献
15.
The presence of ethylenediamine-N-(o-hydroxyphenylacetic)-N'-(p-hydroxyphenylacetic) acid (o,p-EDDHA) as the second largest component in commercial EDDHA iron chelates has recently been demonstrated. Here is reported the speciation of o,p-EDDHA by the application of a novel methodology through the determination of the complexing capacity, protonation, and Ca(2+), Mg(2+), Cu(2+), and Fe(3+) stability constants. The pM values and species distribution in solution, hydroponic, and soil conditions were obtained. Due to the para position of one phenol group in o,p-EDDHA, the protonation constants and Ca and Mg stability constants have different values from those of o,o-EDDHA and p,p-EDDHA regioisomers. o,p-EDDHA/Fe(3+) stability constants are higher than those of EDTA/Fe(3+) but lower than those of o,o-EDDHA/Fe(3+). The sequence obtained for pFe is o,o-EDDHA/Fe(3+) >/= o,p-EDDHA/Fe(3+) > EDTA/Fe(3+). o,p-EDDHA/Fe(3+) can be used as an iron chelate in hydroponic conditions. Also, it can be used in soils with limited Cu availability. 相似文献
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(R)-Terpinen-4-ol was mixed in an artificial diet at a concentration of 1 mg/g of diet, and the diet was fed to the last instar larvae of common cutworm (Spodoptera litura). Metabolites were recovered from frass and analyzed spectroscopically. (R)-Terpinen-4-ol was transformed mainly to (R)-p-menth-1-en-4,7-diol. Similarly, (S)-terpinen-4-ol was transformed mainly to (S)-p-menth-1-en-4,7-diol. The C-7 position (allylic methyl group) of (R)- and (S)-terpinen-4-ol was preferentially oxidized. 相似文献