含LacZ重组逆转录病毒的制备及其在NIH3T3细胞的表达

构建了含有3．7kbLacZ的重组逆转录病毒载体,转染包装细胞系PA317后,经G418筛选,得到了G418的抗性的产毒细胞系,用收获的含有重组逆转录病毒粒子的浓缩病毒上清,感染NIH3T3细胞。经X-gal染色检测,发现表达了LacZ的NIH3T3细胞呈蓝色。     

果实特异启动子调控薄皮甜瓜ACC合成酶cDNA反义表达载体的构建

应用反义RNA技术抑制甜瓜成熟过程中内源乙烯的合成,从而培育耐贮运品种是解决甜瓜延熟保鲜难题的可行新方法。根据GenBank中甜瓜、黄瓜ACC合成酶基因氨基酸保守序列设计引物,从成熟的薄皮甜瓜(齐甜1号)果肉组织中提取总RNA,经RT-PCR扩增得到约0.7kb的ACC合成酶cDNA片段,将其克隆到质粒载体pGEM-TEasy中,测序表明,该基因为777bp,编码258个氨基酸;从番茄(东农706)叶片组织中提取总DNA,经PCR扩增得到约2.2kb的E8基因片段,将其克隆到质粒载体pGEM-TEasy中,测序表明,该基因为2192bp;以pCAM2301为起始植物表达载体,pCAM-GT为中间载体,成功构建了果实特异启动子(E8)调控薄皮甜瓜ACC合成酶cDNA反义表达载体,采用冻融法将其转入根癌农杆菌LBA4404,得到了完整的Ti质粒表达载体系统。     

杨梅酸性转化酶基因cDNA分离及表达分析

杨梅果实富含蔗糖,酸性转化酶是蔗糖代谢关键酶,根据植物酸性转化酶基因保守区序列设计引物,提取杨梅叶片RNA,逆转录获得cDNA,以此为模板通过PCR技术扩增到长度为516bp的基因片段,克隆入pMD18-T载体中,命名为MrIVR1(GenBank:DQ339699)。测序及同源性检索表明,该基因推导氨基酸序列与君子兰、葡萄、草莓、胡萝卜等酸性转化酶基因氨基酸序列同源性为60%～69%。运用ClustalX软件对植物转化酶基因进行了系统树分析,结果显示,MrIVR1编码的蛋白质属于细胞壁酸性转化酶。半定量RT-PCR表达分析显示,MrIVR1基因在杨梅果实发育早期表达量最高,随着果实的发育表达量下降,在成熟果实中表达水平较低。     

Mutated sodium channel genes and elevated monooxygenases are found in pyrethroid resistant populations of Sri Lankan malaria vectors

The present status of pyrethroid resistance in vectors of malaria; Anopheles culicifacies and Anopheles subpictus, was tested in two malarious Districts, Anuradhapura and Trincomalee, of Sri Lanka. Both species were resistant to permethrin and susceptible to cypermethrin and cyfluthrin. An. subpictus were resistant to deltamethrin. λ-Cyhalothrin and etofenprox resistance was shown only by Anuradhapura An. subpictus. Although there were no differences among the populations for esterase and glutathione S-transferase activities, increased monooxygenase levels were found among Trincomalee populations. The voltage-gated sodium channel gene, the target site gene of pyrethroids, was partially sequenced to screen for mutations previously associated with insecticide resistance. The classic leucine to phenylalanine substitution, TTA to TTT, was detected in An. subpictus. It appears that both kdr type and monooxygenase resistance underlie pyrethroid resistance in these two malaria vectors of Sri Lanka.     

Detection and identification of the phytoplasma associated with pear decline in Taiwan

Pear decline (PD) is an important phytoplasmal disease that occurs mainly in Europe and North America. In 1994, pear trees exhibiting symptoms typical of PD disease were observed in orchards of central Taiwan. The sequence of 16S rDNA and 16S–23S rDNA intergenic spacer region (ISR) of the causative agent of pear decline in Taiwan (PDTW) were amplified with polymerase chain reaction (PCR) using a DNA template prepared from the diseased leaves. Sequence analysis of 16S rDNA revealed that the PDTW agent was closely related to the phytoplasmas of the apple proliferation group that cause diseases in stone fruits, pear and apple. Consistent with the result of 16S rDNA sequence analysis, sequence analysis of the 16S–23S rDNA ISR and putative restriction site analyses of 16S rDNA and 16S–23S rDNA ISR sequences provided further support for the view that the PDTW phytoplasma causing pear decline in Taiwan may represent a new subgroup of the apple proliferation group. According to the rDNA sequence of PDTW phytoplasma, two specific PCR primer pairs, APf2/L1n and fPD1/rPDS1, were designed in this study for the detection of the etiological agent in pear trees and insect vectors. Based on the sequence analyses of the PCR-amplified fragments, two species of pear psyllas, Cacopsylla qianli and Cacopsylla chinensis, were found to carry PDTW phytoplasma.     

所有权概念探讨

在对比各国对所有权的不同界定的基础上,立足于我国国情,重新阐释了所有权的概念。     

THE INFLUENCE OF TYLOSES ON THE MANUFACTURE OF MATCHES

This article deals with the results of time studies of pruning P. radiata from 22 ft. to 35 ft. with pole saws. The influence on pruning time of mean diameter and number of branches removed in the pruning operation, number of cones removed and time of day is investigated and a multiple regression equation, expressing the relationship between pruning time and these variables, is computed. A taper table is prepared and used to investigate the effect of pruning on the formation of knot-free timber. The economics of high-pruning is discussed.     

大豆疫霉菌多聚半乳糖醛酸酶pspg1基因的克隆及表达分析

大豆疫病严重影响我同及世界各国的农业生产,为探讨多聚半乳糖醛酸酶在大豆疫霉菌致病过程中的作用,采用PCR的方法从大豆疫霉菌中克隆了多聚半乳糖醛酸酶pspg1基因,并利用RT-PCR法对其在大豆中的表达进行了分析.结果表明:大豆疫霉菌pspg1基因开放阅读框长1236 bp,编码一个长412氨基酸的蛋白质.对其进化关系进行分析,发现该基因与其它卵菌的pg基因亲缘关系最近,形成一个独立的分支.RT-PCR分析表明:pspg1基因在接种大豆疫霉菌的大豆下胚轴中大量表达,而在健康大豆下胚轴中未检测到.克隆了大豆疫霉菌pspg1基因,并发现该基因在大豆疫霉菌侵染大豆过程中发挥重要作用.     

小麦淀粉合酶基因I的克隆及反义和RNAi载体的构建

采用RT-PCR方法从小麦品种豫教2号发育的籽粒中克隆出淀粉合酶I基因(starch synthase I,SSI)部分cDNA片段(795 bp)(GenBank No.EF221762),同源性比较结果显示,它与GenBank上已报道的SSI基因有高度同源性。以p WM101质粒为基础,构建了由35S启动子调控的SS I基因的反义表达载体p WM101SSI;另外,还以pFGC5941质粒为基础,构建了SSI基因的RNAi载体pFGC5941SSIsa,这些载体的构建为研究此基因的功能奠定了基础。