[Objective] This study was to develop a live vector vaccine of goat pox virus of Peste des petits ruminants(PPR). [Method] Using PCR amplification technique, PPR H gene was obtained, then ligated into pGEM-T easy vect...[Objective] This study was to develop a live vector vaccine of goat pox virus of Peste des petits ruminants(PPR). [Method] Using PCR amplification technique, PPR H gene was obtained, then ligated into pGEM-T easy vector; the recombinants were digested by Nhe Ⅰ and Hind Ⅲ, and ligated into pEGFP-N1-P7.5, yielding the recombinant vector pEGFP-N1-P7.5-H; next the expression cassette EGFP-N1-P7.5-H was first released from recombinant vector pEGFP-N1-P7.5-H by double digestion of Hind Ⅲ and Nhe Ⅰ and ligated into pUC119-TK that was digested by Kpn Ⅰ, yielding the transfer vector pUC119-TK-EGFP-P7.5-H. [Result] Identification and double enzyme digestion showed that the transfer vector pUC119-TK-EGFP-P7.5-H was correctly constructed. From the transfer vector transfected BHK-21 cells which infected GTPV AV41, specific fluorescence was observed at 48th h of transfection. [Conclusion] The construction of goat poxvirus live vector laid a foundation for the live vector vaccine of PPR vaccine.展开更多
利用RACE(rapid-amplification of cDNA end)方法,以大豆叶片提取总的RNA为模板克隆了大豆Na+/H+逆向转运蛋白(Glycine max Na+/H+antiporter,GmNHX)基因,并将其连接到表达载体PBI121中,构建重组表达载体PBI121-NHX3.分析结果表明:该基...利用RACE(rapid-amplification of cDNA end)方法,以大豆叶片提取总的RNA为模板克隆了大豆Na+/H+逆向转运蛋白(Glycine max Na+/H+antiporter,GmNHX)基因,并将其连接到表达载体PBI121中,构建重组表达载体PBI121-NHX3.分析结果表明:该基因的ORF为1 503 bp,推测编码501个氨基酸.与所选取的10种植物同类蛋白氨基酸序列进行对比,一致性为72%~94%,并具有真核生物单价阳离子(氢离子)反向转运蛋白典型的结构域,将该基因命名为GmNHX3,GenBank接收号为JN872904.通过PCR和酶切鉴定,PBI121-NHX3构建成功.展开更多
cDNA libraries were constructed from the leaves of a rice (Oryza sativa L.) salt tolerancevariety Tesan抋i 2 growing in solutions with 150 mmol/L NaCl for 3 h or without salt stress. Three salt-responsive cDNA clones,...cDNA libraries were constructed from the leaves of a rice (Oryza sativa L.) salt tolerancevariety Tesan抋i 2 growing in solutions with 150 mmol/L NaCl for 3 h or without salt stress. Three salt-responsive cDNA clones, Ts1, Ts2 and Ts3 were isolated by differential screening. Northern blottinganalysis showed that the transcription levels of Ts1 and Ts2 increased within 3 h salt stress and kept onincreasing within 24 h, while the transcription level of Ts3 reached its peak within 3 h. Sequence analysisindicated that there were no homologies between the three cDNA clones and any known gene. The threecDNA clones were mapped using a doubled haploid (DH) population derived from an indica variety ZYQ8,which was a salt tolerance parent of Tesan抋i 2, with a japonica variety JX17. Ts1, Ts2 and Ts3 werelocated on chromosomes 1, 3 and 7, respectively. It was noted that Ts1, Ts2, and Ts3 were in or near theregions of major or minor salt tolerance quantitative trait loci (QTLs), which were mapped in the same DHpopulation in a parallel study.展开更多
文摘[Objective] This study was to develop a live vector vaccine of goat pox virus of Peste des petits ruminants(PPR). [Method] Using PCR amplification technique, PPR H gene was obtained, then ligated into pGEM-T easy vector; the recombinants were digested by Nhe Ⅰ and Hind Ⅲ, and ligated into pEGFP-N1-P7.5, yielding the recombinant vector pEGFP-N1-P7.5-H; next the expression cassette EGFP-N1-P7.5-H was first released from recombinant vector pEGFP-N1-P7.5-H by double digestion of Hind Ⅲ and Nhe Ⅰ and ligated into pUC119-TK that was digested by Kpn Ⅰ, yielding the transfer vector pUC119-TK-EGFP-P7.5-H. [Result] Identification and double enzyme digestion showed that the transfer vector pUC119-TK-EGFP-P7.5-H was correctly constructed. From the transfer vector transfected BHK-21 cells which infected GTPV AV41, specific fluorescence was observed at 48th h of transfection. [Conclusion] The construction of goat poxvirus live vector laid a foundation for the live vector vaccine of PPR vaccine.
文摘利用RACE(rapid-amplification of cDNA end)方法,以大豆叶片提取总的RNA为模板克隆了大豆Na+/H+逆向转运蛋白(Glycine max Na+/H+antiporter,GmNHX)基因,并将其连接到表达载体PBI121中,构建重组表达载体PBI121-NHX3.分析结果表明:该基因的ORF为1 503 bp,推测编码501个氨基酸.与所选取的10种植物同类蛋白氨基酸序列进行对比,一致性为72%~94%,并具有真核生物单价阳离子(氢离子)反向转运蛋白典型的结构域,将该基因命名为GmNHX3,GenBank接收号为JN872904.通过PCR和酶切鉴定,PBI121-NHX3构建成功.
文摘cDNA libraries were constructed from the leaves of a rice (Oryza sativa L.) salt tolerancevariety Tesan抋i 2 growing in solutions with 150 mmol/L NaCl for 3 h or without salt stress. Three salt-responsive cDNA clones, Ts1, Ts2 and Ts3 were isolated by differential screening. Northern blottinganalysis showed that the transcription levels of Ts1 and Ts2 increased within 3 h salt stress and kept onincreasing within 24 h, while the transcription level of Ts3 reached its peak within 3 h. Sequence analysisindicated that there were no homologies between the three cDNA clones and any known gene. The threecDNA clones were mapped using a doubled haploid (DH) population derived from an indica variety ZYQ8,which was a salt tolerance parent of Tesan抋i 2, with a japonica variety JX17. Ts1, Ts2 and Ts3 werelocated on chromosomes 1, 3 and 7, respectively. It was noted that Ts1, Ts2, and Ts3 were in or near theregions of major or minor salt tolerance quantitative trait loci (QTLs), which were mapped in the same DHpopulation in a parallel study.