Establishment of a tetracycline-off and heat shock-on gene expression system in tobacco  

Establishment of a tetracycline-off and heat shock-on gene expression system in tobacco

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作  者:ZHOU You LI Jin-hua PAN Yu ZHENG Yu PAN Yang-lu DING Yu-mei SU Cheng-gang ZHANG Xing-guo 

机构地区:[1]Key Laboratory of Horticulture Science for Southern Mountainous Regions,Ministry of Education/College of Horticulture and Landscape Architecture,Southwest University,Chongqing 400715,P.R.China

出  处:《Journal of Integrative Agriculture》2017年第5期1112-1119,共8页农业科学学报(英文版)

基  金:supported by the National High-Tech R&D Program of China(2010AA10060705);the Transgenic Engineering Crops Breeding Special Funds from China’s Ministry of Agriculture(2009ZX08010-005B)

摘  要:The tetracycline (Tet)-off gene expression regulation system based on the TetR-VP16/Top10 construct has not been widely utilized in plants, for its highly expressed TetR-VP16 activator is toxic to some plants and repeatedly replenishing tetracycline to turn off the constitutively active system is a tedious process. To solve these problems, a Tet-off and heat shock (HS)-on gene expression regulation system was constructed in this study. This system is composed of a chimeric transactivator gene TetR-HSF that is derived from a Tet repressor (TetR) and a HS transcription factor (HSF) controlled by a HS promoter HSP70m, and a Tet operator containing hybrid promoter, Om35S, that drives expression of the β-glucuronidase (GUS) gene. The resultant system yields a GUS expression pattern similar to that of the HSP70m promoter under inducing temperatures and at 35 and 40℃ drives GUS expression to a similar level as the Cauliflower mosaic virus (CaMV) 35S promoter. Further examination revealed that the TetR-HSF and GUS genes were induced by HS, reaching peak expression after 1 and 6 h treatment, respectively, and the HS induction of the expression system could be inhibited by Tet. This system will provide a useful tool for transgenic studies of plants in the laboratory and in the field, including transgene function analysis, agronomic trait improvement, biopharmaceutical protein production and others.The tetracycline (Tet)-off gene expression regulation system based on the TetR-VP16/Top10 construct has not been widely utilized in plants, for its highly expressed TetR-VP16 activator is toxic to some plants and repeatedly replenishing tetracycline to turn off the constitutively active system is a tedious process. To solve these problems, a Tet-off and heat shock (HS)-on gene expression regulation system was constructed in this study. This system is composed of a chimeric transactivator gene TetR-HSF that is derived from a Tet repressor (TetR) and a HS transcription factor (HSF) controlled by a HS promoter HSP70m, and a Tet operator containing hybrid promoter, Om35S, that drives expression of the β-glucuronidase (GUS) gene. The resultant system yields a GUS expression pattern similar to that of the HSP70m promoter under inducing temperatures and at 35 and 40℃ drives GUS expression to a similar level as the Cauliflower mosaic virus (CaMV) 35S promoter. Further examination revealed that the TetR-HSF and GUS genes were induced by HS, reaching peak expression after 1 and 6 h treatment, respectively, and the HS induction of the expression system could be inhibited by Tet. This system will provide a useful tool for transgenic studies of plants in the laboratory and in the field, including transgene function analysis, agronomic trait improvement, biopharmaceutical protein production and others.

关 键 词:gene expression system heat shock-on tetracycline-off TetR-HSF transactivator TOBACCO 

分 类 号:Q943.2[生物学—植物学]

 

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