The receptor for activated C-kinase 1 （RACK1） is a highly conserved scaffold protein with versatile functions, and plays important roles in the regulation of plant growth and development. Transgenic rice plants, in ...The receptor for activated C-kinase 1 （RACK1） is a highly conserved scaffold protein with versatile functions, and plays important roles in the regulation of plant growth and development. Transgenic rice plants, in which the expression of RACK1 gene was inhibited by RNA interference （RNAi）, were studied to elucidate the possible functions of RACK1 in responses to drought stress in rice. Real-time PCR analysis showed that the expression of RACK1 in transgenic rice plants was inhibited by more than 50%. The tolerance to drought stress of the transgenic rice plants was higher as compared with the non-transgenic rice plants. The peroxidation of membrane and the production of malondialdehyde were significantly lower and the superoxide dismutase activity in transgenic rice plants was significantly higher than those in non-trangenic rice plants It is suggested that RACK1 negatively regulated the redox system-related tolerance to drought stress of rice plants.展开更多
The Citrus tristeza virus (CTV) uses 3 silencing suppressor genes, p20, p23 and p25, to resist the attacks from its Citrus hosts. Inactivating these genes is therefore obviously a potential defensive option in additio...The Citrus tristeza virus (CTV) uses 3 silencing suppressor genes, p20, p23 and p25, to resist the attacks from its Citrus hosts. Inactivating these genes is therefore obviously a potential defensive option in addition to the current control strat-egies including aphid management and the use of mild strain cross protection. In this study, we cloned partial DNA frag-ments from the three genes, and used them to construct vectors for expressing hairpin RNAs (hpRNAs). To facilitate the formation of hpRNAs, the constructs were introduced in a loop structure. Fol owing transformation of sour orange (Citrus aurantium) with these constructs, 8 p20 hpRNA (hp20) and 1 p25 hpRNA (hp25) expressing lines were obtained. The 7 hp20 transgenic lines were further characterized. Their reactions to CTV were tested fol owing inoculation with CT14A and/or TR-L514, both of which are severe strains. Results showed that 3 lines (hp20-5, hp20-6 and hp20-8) were completely resistant to TR-L514 under greenhouse conditions for no detectable viral load was found in their leaves by PCR. However, they exhibited only partial suppression of TR-L514 under screen house conditions since the virus was detected in their leaves, though 2 months later compared to non-transgenic controls. Further tests showed that hp20-5 was tolerant also to CT14A under screen house conditions. The growth of hp20-5 was much better than others including the controls that were concurrently chal enged with CT14A. These results showed that expressing p20 hpRNA was sufifcient to confer sour orange with CTV resistance/tolerance.展开更多
目的:探讨腺病毒介导的KDR启动子驱动的CD/TK双自杀基因系统(Ad-KDRP-CD/TK)联合survivin基因干扰对肝癌细胞在裸鼠体内生长的抑制作用。方法:将20只裸鼠随机分均为模型组(皮下植入BEL-7402肝癌细胞成瘤,不加任何处理)、双自杀基因转染...目的:探讨腺病毒介导的KDR启动子驱动的CD/TK双自杀基因系统(Ad-KDRP-CD/TK)联合survivin基因干扰对肝癌细胞在裸鼠体内生长的抑制作用。方法:将20只裸鼠随机分均为模型组(皮下植入BEL-7402肝癌细胞成瘤,不加任何处理)、双自杀基因转染组(皮下植入转染Ad-KDRP-CD/TK的BEL-7402细胞,成瘤后瘤内注射前药更昔洛韦与5-氟胞嘧啶)、survivin si RNA转染组(皮下注射BEL-7402肝癌细胞,成瘤后瘤内注射survivin si RNA/LipDMEM转染混合物)、联合转染组(双自杀基因转染+survivin si RNA转染处理)。治疗2周后处死各组小鼠,称取肿瘤质量,计算肿瘤抑制率,检测瘤组织微血管密度(MVD)及survivin m RNA与蛋白表达。结果:各治疗组的肿瘤质量明显小于模型组(均P<0.05),且联合转染组的抑瘤率最大(均P<0.05);肿瘤组织MVD、survivin m RNA与蛋白水平均明显降低,且联合转染组的降低程度最为明显(均P<0.05)。结论:双自杀基因联合survivin干扰是抑制鼠肝癌细胞在体内生长的有效途径。展开更多
Silencing of gene expression by RNA interference (RNAi) has become a widely used tool. For the study of mammalian gene function expression vectors for short hairpin RNA (shRNA) were developed. However the standard met...Silencing of gene expression by RNA interference (RNAi) has become a widely used tool. For the study of mammalian gene function expression vectors for short hairpin RNA (shRNA) were developed. However the standard methods of shRNA transgenic (Tg) mice production have not been established. Sry (sex-determining region on the Y chromosome) is a mammalian sex-determining gene on the Y chromosome. In mice, the transient expression of Sry in supporting cell precursor cells between 10.5 and 12.5 days post-coitus (dpc) triggers the differentiation of Sertoli cells from granulosa cells. Then high efficiency of Sry gene silencing in Tg mice should induce XY male-to-female sex reversal. An shRNA Tg mouse targeting Sry gene was attempted to be generated by pronuclear microinjection. A low rate (Tg pups/all pups born after microinjection = 2/154 to 7/178) of Tg pups was observed. These Tg mice showed no XY male-to-female sex reversal. The results suggest that exogenous expression of small RNA might exert a negative effect on embryonic development and another approach should be needed for RNAi transgenesis in mice.展开更多
基金supported by the National Natural Science Foundation of China (Grant No. 30571120)the National High Technology Research and Development Program of China (Grant No.2008AA10Z120)the Research Fund for the Doctoral Program of Higher Education, China
文摘The receptor for activated C-kinase 1 （RACK1） is a highly conserved scaffold protein with versatile functions, and plays important roles in the regulation of plant growth and development. Transgenic rice plants, in which the expression of RACK1 gene was inhibited by RNA interference （RNAi）, were studied to elucidate the possible functions of RACK1 in responses to drought stress in rice. Real-time PCR analysis showed that the expression of RACK1 in transgenic rice plants was inhibited by more than 50%. The tolerance to drought stress of the transgenic rice plants was higher as compared with the non-transgenic rice plants. The peroxidation of membrane and the production of malondialdehyde were significantly lower and the superoxide dismutase activity in transgenic rice plants was significantly higher than those in non-trangenic rice plants It is suggested that RACK1 negatively regulated the redox system-related tolerance to drought stress of rice plants.
基金supported by the International Science & Technology Cooperation Program of China (2012DFA30610)the National Natural Science Foundation of China (30571291)the Special Fund for Agro-Scientific Research in the Public Interest, China (201203075-07)
文摘The Citrus tristeza virus (CTV) uses 3 silencing suppressor genes, p20, p23 and p25, to resist the attacks from its Citrus hosts. Inactivating these genes is therefore obviously a potential defensive option in addition to the current control strat-egies including aphid management and the use of mild strain cross protection. In this study, we cloned partial DNA frag-ments from the three genes, and used them to construct vectors for expressing hairpin RNAs (hpRNAs). To facilitate the formation of hpRNAs, the constructs were introduced in a loop structure. Fol owing transformation of sour orange (Citrus aurantium) with these constructs, 8 p20 hpRNA (hp20) and 1 p25 hpRNA (hp25) expressing lines were obtained. The 7 hp20 transgenic lines were further characterized. Their reactions to CTV were tested fol owing inoculation with CT14A and/or TR-L514, both of which are severe strains. Results showed that 3 lines (hp20-5, hp20-6 and hp20-8) were completely resistant to TR-L514 under greenhouse conditions for no detectable viral load was found in their leaves by PCR. However, they exhibited only partial suppression of TR-L514 under screen house conditions since the virus was detected in their leaves, though 2 months later compared to non-transgenic controls. Further tests showed that hp20-5 was tolerant also to CT14A under screen house conditions. The growth of hp20-5 was much better than others including the controls that were concurrently chal enged with CT14A. These results showed that expressing p20 hpRNA was sufifcient to confer sour orange with CTV resistance/tolerance.
文摘目的:探讨腺病毒介导的KDR启动子驱动的CD/TK双自杀基因系统(Ad-KDRP-CD/TK)联合survivin基因干扰对肝癌细胞在裸鼠体内生长的抑制作用。方法:将20只裸鼠随机分均为模型组(皮下植入BEL-7402肝癌细胞成瘤,不加任何处理)、双自杀基因转染组(皮下植入转染Ad-KDRP-CD/TK的BEL-7402细胞,成瘤后瘤内注射前药更昔洛韦与5-氟胞嘧啶)、survivin si RNA转染组(皮下注射BEL-7402肝癌细胞,成瘤后瘤内注射survivin si RNA/LipDMEM转染混合物)、联合转染组(双自杀基因转染+survivin si RNA转染处理)。治疗2周后处死各组小鼠,称取肿瘤质量,计算肿瘤抑制率,检测瘤组织微血管密度(MVD)及survivin m RNA与蛋白表达。结果:各治疗组的肿瘤质量明显小于模型组(均P<0.05),且联合转染组的抑瘤率最大(均P<0.05);肿瘤组织MVD、survivin m RNA与蛋白水平均明显降低,且联合转染组的降低程度最为明显(均P<0.05)。结论:双自杀基因联合survivin干扰是抑制鼠肝癌细胞在体内生长的有效途径。
文摘Silencing of gene expression by RNA interference (RNAi) has become a widely used tool. For the study of mammalian gene function expression vectors for short hairpin RNA (shRNA) were developed. However the standard methods of shRNA transgenic (Tg) mice production have not been established. Sry (sex-determining region on the Y chromosome) is a mammalian sex-determining gene on the Y chromosome. In mice, the transient expression of Sry in supporting cell precursor cells between 10.5 and 12.5 days post-coitus (dpc) triggers the differentiation of Sertoli cells from granulosa cells. Then high efficiency of Sry gene silencing in Tg mice should induce XY male-to-female sex reversal. An shRNA Tg mouse targeting Sry gene was attempted to be generated by pronuclear microinjection. A low rate (Tg pups/all pups born after microinjection = 2/154 to 7/178) of Tg pups was observed. These Tg mice showed no XY male-to-female sex reversal. The results suggest that exogenous expression of small RNA might exert a negative effect on embryonic development and another approach should be needed for RNAi transgenesis in mice.
