西花蓟马酵母双杂交文库及TSWV膜蛋白诱饵载体的构建
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摘要
【目的】为筛选西花蓟马Frankliniella occidentalis与番茄斑萎病毒的互作蛋白。【方法】利用MakeYour Own"Mate&Plate?"Library System构建西花蓟马酵母双杂交初级cDNA文库以及Y187酵母次级文库,利用Matchmaker Gold Yeast Two-Hybrid System构建TSWV膜蛋白GN及GC诱饵载体。【结果】建立的西花蓟马初级cDNA文库库容为5.6×106 cfu,其插入片段平均长度大于1 000 bp,重组率约为100%。建立的Y187酵母次级文库转化效率为5×106cfu/μg,文库滴度为2.97×108。构建的pGNKT7-GN及pGBKT7-GC诱饵载体能够在Y2HGold酵母菌株中正确表达,无自激活活性且无毒性。【结论】成功构建了西花蓟马酵母双杂交文库以及番茄斑萎病毒膜蛋白诱饵载体,可用于后续筛库实验。
[Objectives] To screen proteins involved in the interaction between western flower thrips and the Tomato spotted wilt virus(TSWV). [Methods] A western flower thrip cDNA library and a Y187 yeast library were constructed using the Make Your Own "Mate & Plate?" Library System, and TSWV GN and GC bait vectors were constructed using the Matchmaker Gold Yeast Two-Hybrid System. [Results] The capacity of the western flower thrip cDNA library was 5.6×106 cfu,the average library recombination rate was about 100%, and the average amplification sizes of insert fragments in the cDNA library were above 1 kb. The transformation ratio and titer of the Y187 yeast library were 5×106 cfu/μg and 2.97×108,respectively. The constructed pGNKT7-GN and pGBKT7-GC bait vectors were well expressed in yeast and had no self-activation and toxicity. [Conclusion] A western flower thrip yeast library and TSWV membrane protein bait vectors were successfully constructed and can be used for further hybrid library screening.
[Objectives] To screen proteins involved in the interaction between western flower thrips and the Tomato spotted wilt virus(TSWV). [Methods] A western flower thrip cDNA library and a Y187 yeast library were constructed using the Make Your Own "Mate & Plate?" Library System, and TSWV GN and GC bait vectors were constructed using the Matchmaker Gold Yeast Two-Hybrid System. [Results] The capacity of the western flower thrip cDNA library was 5.6×106 cfu,the average library recombination rate was about 100%, and the average amplification sizes of insert fragments in the cDNA library were above 1 kb. The transformation ratio and titer of the Y187 yeast library were 5×106 cfu/μg and 2.97×108,respectively. The constructed pGNKT7-GN and pGBKT7-GC bait vectors were well expressed in yeast and had no self-activation and toxicity. [Conclusion] A western flower thrip yeast library and TSWV membrane protein bait vectors were successfully constructed and can be used for further hybrid library screening.
引文
Adkins S,2000.Tomato spotted wilt virus positive steps towards negative success.Molecular Plant Pathology,1(3):151-157.
Fields S,Song O,1989.A novel genetic system to detect proteinprotein interactions.Nature,340:245-246.
Jones DR,2005.Plant viruses transmitted by thrips.European Journal of Plant Pathology,113(2):119-157.
Kikkert M,Meurs C,van de Wetering F,Dorfmuller S,Peters D,Kormelink R,Goldbach R,1998.Binding of tomato spotted wilt virus to a 94-kDa thrips protein.Phytopathology,88(1):63-69.
Kirk WDJ,Terry LI,2003.The spread of the western flower thrips Frankliniella occidentalis(Pergande).Agricultural and Forest Entomology,5(4):301-310.
Lewis T,1997.Thrips as Crop Pests.London:Cambridge Press.1-13.
Li S,Xiong RY,Wang XF,Zhou YJ,2011.Five proteins of Laodelphax striatellus are potentially involved in the interactions between rice stripe virus and vector.PLoS ONE,6(10):e26585.
Lin L,Shi YH,Luo ZP,Lu YW,Zheng HY,Yan F,Chen J,Chen JP,Adams MJ,Wu YF,2009.Protein-protein interactions in two potyviruses using the yeast two-hybrid system.Virus Research,142(1/2):36-40.
Medeiros RB,Ullman DE,Sherwood JL,German TL,2000.Immunoprecipitation of a 50-kDa protein:a candidate receptor component for a tomato spotted wilt tospovirus(Bunyaviridae)in its main vector,Franklinella occidentalis.Virus Research,67(2):109-118.
Montero-Astúa M,Rotenberg D,Leach-Kieffaber A,Schneweis BA,Park S,Park JK,German TL,Whitfield AE,2014.Disruption of vector transmission by a plant-expressed viral glycoprotein.Molecular Plant-Microbe Interactions,27(3):296-304.
Morin S,Ghanim M,Sobol I,Czosnek H,2000.The Gro EL protein of the whitefly Bemisia tabaci interacts with the coat protein of transmissible and nontransmissible begomoviruses in the yeast two-hybrid system.Virology,276(2):404-416.
Moritz G,Kumm S,Mound L,2004.Tosporirus transmission depends on thrips ontogeny.Virus Research,100(1):143-149.
Nagata T,Inoue-Nagata AK,Prins M,Goldbach R,Peters D,2000.Impeded thrips transmission of defective Tomato spotted wilt virus isolates.Phytopathology,90(5):454-459.
Scholthof KBG,Adkins S,Czosnek H,Palukaitis P,Jacquot E,Hohn T,Hohn B,Saunders K,Candresse T,Ahlquist P,Hemenway C,Foster GD,2011.Top 10 plant virus in molecular plant pathology.Molecular Plant Pathology,12(9):938-954.
Seo JK,Hwang SH,Kang SH,Choi HS,Lee SH,Sohn SH,Kim KH,2007.Interaction study of soybean mosaic virus proteins with soybean proteins using the yeast-two hybrid system.The Plant Pathology Journal,23(4):281-286.
Ullman DE,Medeiros RB,Campbell LR,Whitfield AE,Sherwood JL,German TL,2002.Thrips as vectors of tospoviruses.Advances in Botanical Research,36(1):113-140.
van de Wetering F,Goldbach R,Peters D,1996.Tomato spotted wilt tospovirus ingestion by first instar larvae of Frankliniella occidentalis is a prerequisite for transmission.Phytopathology,86(9):900-905.
Wang HH,Lei ZR,Li X,Dai AG,Chen HQ,2013.An important invasive pest,Frankliniella occidentalis,inspected in Tibet.Plant Protection,39(1):181-183.[王海鸿,雷仲仁,李雪,代安国,陈翰秋,2013.西藏发现重要外来入侵害虫--西花蓟马.植物保护,39(1):181-183.]
Whitfield AE,Ullman DE,German TL,2004.Expression and characterization of a soluble form of tomato spotted wilt virus glycoprotein GN.Journal of Virology,78(23):13197-131206.
Whitfield,AE,Ullman DE,German TL,2005.Tospovirus-thrips interactions.Annual Review of Phytopathology,43(1):459-489.
Wijkamp I,van Lent J,Kormelink R,Kormelink R,Godbach R,Peters D,1993.Multiplication of tomato spotted wilt virus in its insect vector,Frankliniella occidentalis.Journal of General Virology,74(3):341-349.
Wu QJ,Xu BY,Zhang ZJ,Zhang YJ,Zhu GR,2007.Distribution and species of thrips in Beijing,Zhejiang,Yunnan region.China Plant Protection,27(1):32-34.[吴青君,徐宝云,张治军,张友军,朱国仁,2007.京、浙、滇地区植物蓟马种类及其分布调查.中国植保导刊,27(1):32-34.]
Yuan CM,Zhi JR,Li JZ,Zhang Y,2008.The distribution and integrated management of vegetables thrips in Guizhou Province.Hubei Agricultural Science,47(12):1442-1444.[袁成明,郅军锐,李景柱,张勇,2008.贵州省蔬菜蓟马的种类、分布及综合防治.湖北农业科学,47(12):1442-1444.]
Zhang YJ,Wu QJ,Xu BY,Zhu GR,2003.Dangerous invasive pestFrankliniella occidentalis occurred in Beijing.Plant Protection,29(4):58-59.[张友军,吴青君,徐宝云,朱国仁,2003.危险性外来入侵生物-西花蓟马在北京发生危害.植物保护,29(4):58-59.]
Zhang YJ,Zhu GR,Chu D,Wu QJ,Wang SL,2011.Occurrence,damage and control of important invasive insect pests on the vegetable crops in China.Plant Protection,37(4):1-6.[张友军,朱国仁,褚栋,吴青君,王少丽,2011.我国蔬菜作物重大入侵害虫发生、危害与控制.植物保护,37(6):1-6.]
Zhang ZJ,Zhang PJ,Li WD,Zhang JM,Huang F,Yang J,Bei YW,Lu YB,2013.De novo transcriptome sequencing in Frankliniella occidentalis to identify genes involved in plant virus transmission and insecticide resistance.Genomics,101(5):296-305.
Zheng CY,Liu YH,Zhang NQ,Zhao XL,2007.Invaded insect pest-Frankliniella occidentalis first reported in Shandong province.Journal of Qingdao Agricultural University(Natural Science),24(3):l72-174.[郑长英,刘云虹,张乃芹,赵希丽,2007.山东省发现外来入侵有害生物-西花蓟马.青岛农业大学学报(自然科学版),24(3):l72-174.]
Fields S,Song O,1989.A novel genetic system to detect proteinprotein interactions.Nature,340:245-246.
Jones DR,2005.Plant viruses transmitted by thrips.European Journal of Plant Pathology,113(2):119-157.
Kikkert M,Meurs C,van de Wetering F,Dorfmuller S,Peters D,Kormelink R,Goldbach R,1998.Binding of tomato spotted wilt virus to a 94-kDa thrips protein.Phytopathology,88(1):63-69.
Kirk WDJ,Terry LI,2003.The spread of the western flower thrips Frankliniella occidentalis(Pergande).Agricultural and Forest Entomology,5(4):301-310.
Lewis T,1997.Thrips as Crop Pests.London:Cambridge Press.1-13.
Li S,Xiong RY,Wang XF,Zhou YJ,2011.Five proteins of Laodelphax striatellus are potentially involved in the interactions between rice stripe virus and vector.PLoS ONE,6(10):e26585.
Lin L,Shi YH,Luo ZP,Lu YW,Zheng HY,Yan F,Chen J,Chen JP,Adams MJ,Wu YF,2009.Protein-protein interactions in two potyviruses using the yeast two-hybrid system.Virus Research,142(1/2):36-40.
Medeiros RB,Ullman DE,Sherwood JL,German TL,2000.Immunoprecipitation of a 50-kDa protein:a candidate receptor component for a tomato spotted wilt tospovirus(Bunyaviridae)in its main vector,Franklinella occidentalis.Virus Research,67(2):109-118.
Montero-Astúa M,Rotenberg D,Leach-Kieffaber A,Schneweis BA,Park S,Park JK,German TL,Whitfield AE,2014.Disruption of vector transmission by a plant-expressed viral glycoprotein.Molecular Plant-Microbe Interactions,27(3):296-304.
Morin S,Ghanim M,Sobol I,Czosnek H,2000.The Gro EL protein of the whitefly Bemisia tabaci interacts with the coat protein of transmissible and nontransmissible begomoviruses in the yeast two-hybrid system.Virology,276(2):404-416.
Moritz G,Kumm S,Mound L,2004.Tosporirus transmission depends on thrips ontogeny.Virus Research,100(1):143-149.
Nagata T,Inoue-Nagata AK,Prins M,Goldbach R,Peters D,2000.Impeded thrips transmission of defective Tomato spotted wilt virus isolates.Phytopathology,90(5):454-459.
Scholthof KBG,Adkins S,Czosnek H,Palukaitis P,Jacquot E,Hohn T,Hohn B,Saunders K,Candresse T,Ahlquist P,Hemenway C,Foster GD,2011.Top 10 plant virus in molecular plant pathology.Molecular Plant Pathology,12(9):938-954.
Seo JK,Hwang SH,Kang SH,Choi HS,Lee SH,Sohn SH,Kim KH,2007.Interaction study of soybean mosaic virus proteins with soybean proteins using the yeast-two hybrid system.The Plant Pathology Journal,23(4):281-286.
Ullman DE,Medeiros RB,Campbell LR,Whitfield AE,Sherwood JL,German TL,2002.Thrips as vectors of tospoviruses.Advances in Botanical Research,36(1):113-140.
van de Wetering F,Goldbach R,Peters D,1996.Tomato spotted wilt tospovirus ingestion by first instar larvae of Frankliniella occidentalis is a prerequisite for transmission.Phytopathology,86(9):900-905.
Wang HH,Lei ZR,Li X,Dai AG,Chen HQ,2013.An important invasive pest,Frankliniella occidentalis,inspected in Tibet.Plant Protection,39(1):181-183.[王海鸿,雷仲仁,李雪,代安国,陈翰秋,2013.西藏发现重要外来入侵害虫--西花蓟马.植物保护,39(1):181-183.]
Whitfield AE,Ullman DE,German TL,2004.Expression and characterization of a soluble form of tomato spotted wilt virus glycoprotein GN.Journal of Virology,78(23):13197-131206.
Whitfield,AE,Ullman DE,German TL,2005.Tospovirus-thrips interactions.Annual Review of Phytopathology,43(1):459-489.
Wijkamp I,van Lent J,Kormelink R,Kormelink R,Godbach R,Peters D,1993.Multiplication of tomato spotted wilt virus in its insect vector,Frankliniella occidentalis.Journal of General Virology,74(3):341-349.
Wu QJ,Xu BY,Zhang ZJ,Zhang YJ,Zhu GR,2007.Distribution and species of thrips in Beijing,Zhejiang,Yunnan region.China Plant Protection,27(1):32-34.[吴青君,徐宝云,张治军,张友军,朱国仁,2007.京、浙、滇地区植物蓟马种类及其分布调查.中国植保导刊,27(1):32-34.]
Yuan CM,Zhi JR,Li JZ,Zhang Y,2008.The distribution and integrated management of vegetables thrips in Guizhou Province.Hubei Agricultural Science,47(12):1442-1444.[袁成明,郅军锐,李景柱,张勇,2008.贵州省蔬菜蓟马的种类、分布及综合防治.湖北农业科学,47(12):1442-1444.]
Zhang YJ,Wu QJ,Xu BY,Zhu GR,2003.Dangerous invasive pestFrankliniella occidentalis occurred in Beijing.Plant Protection,29(4):58-59.[张友军,吴青君,徐宝云,朱国仁,2003.危险性外来入侵生物-西花蓟马在北京发生危害.植物保护,29(4):58-59.]
Zhang YJ,Zhu GR,Chu D,Wu QJ,Wang SL,2011.Occurrence,damage and control of important invasive insect pests on the vegetable crops in China.Plant Protection,37(4):1-6.[张友军,朱国仁,褚栋,吴青君,王少丽,2011.我国蔬菜作物重大入侵害虫发生、危害与控制.植物保护,37(6):1-6.]
Zhang ZJ,Zhang PJ,Li WD,Zhang JM,Huang F,Yang J,Bei YW,Lu YB,2013.De novo transcriptome sequencing in Frankliniella occidentalis to identify genes involved in plant virus transmission and insecticide resistance.Genomics,101(5):296-305.
Zheng CY,Liu YH,Zhang NQ,Zhao XL,2007.Invaded insect pest-Frankliniella occidentalis first reported in Shandong province.Journal of Qingdao Agricultural University(Natural Science),24(3):l72-174.[郑长英,刘云虹,张乃芹,赵希丽,2007.山东省发现外来入侵有害生物-西花蓟马.青岛农业大学学报(自然科学版),24(3):l72-174.]