本文關(guān)鍵詞：番茄斑萎病毒和水稻黑條矮縮病毒CP和RdRp的轉(zhuǎn)基因煙草構(gòu)建　出處：《山東農(nóng)業(yè)大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 反向遺傳學(xué)體系 番茄斑萎病毒 水稻黑條矮縮病毒 CP RdRp 轉(zhuǎn)基因

【摘要】：水稻黑條矮縮病毒(Rice black-streaked dwarf virus,RBSDV)可侵染水稻、玉米、小麥等禾本科作物,引起水稻黑條矮縮病、玉米粗縮病等病害。其基因組有10條雙鏈RNA(S1-S10),S1編碼RNA依賴的RNA聚合酶(RdRp),S10編碼外殼蛋白(CP)。番茄斑萎病毒(Tomato spotted wilt virus,TSWV)能夠侵染70科900多種植物,病害在世界范圍內(nèi)都有分布。其基因組包括三條單鏈RNA(L、M和S);RNA L為負(fù)義RNA,其互補(bǔ)鏈編碼RNA依賴的RNA聚合酶(RdRp),RNA S為雙義RNA,其互補(bǔ)鏈編碼核衣殼蛋白(N)。這兩種病害均可造成嚴(yán)重的生產(chǎn)危害和經(jīng)濟(jì)損失,為進(jìn)一步對這兩種RNA病毒進(jìn)行分子生物學(xué)水平研究,本研究旨在構(gòu)建含有TSWV和RBSDV CP和RdRp蛋白的轉(zhuǎn)基因煙草,嘗試建立其反向遺傳學(xué)體系�；陔p元載體pROKⅡ,構(gòu)建了分別含有RBSDV和TSWV CP和RdRp的pROKⅡ-TS-CP、pROKⅡ-TS-CP-8H、pROKⅡ-RB-CP、pROKⅡ-RB-CP-8H、pROKⅡ-TSRdRp、pROKⅡ-TS-RdRp-8H、pROKⅡ-RB-RdRp、pROKⅡ-RB-RdRp-8H等8個(gè)不帶His標(biāo)簽或帶His標(biāo)簽的轉(zhuǎn)基因載體。采用農(nóng)桿菌介導(dǎo)的葉盤法分別將8個(gè)載體轉(zhuǎn)化普通煙(Nicotiana tabacum,NC89),普通煙葉片經(jīng)過預(yù)培養(yǎng)、共培養(yǎng)、選擇培養(yǎng),篩選出抗性愈傷組織,將獲得不定芽轉(zhuǎn)入生根培養(yǎng)基獲得完整的再生植株。通過DNA、RNA及蛋白水平檢測表明,已得到含有TSWV CP(帶有His標(biāo)簽)和RBSDV CP(帶有His標(biāo)簽)的單價(jià)普通煙,其他單、雙價(jià)轉(zhuǎn)基因煙草正在培養(yǎng)和生長中。
[Abstract]:Rice black-streaked dwarf virus (RBSDV) can infect gramineous crops such as rice, corn, wheat and so on. The genome contains 10 double-stranded RNAs S1-S10 / S1 encoding RNA dependent RNA polymerase (RdRp). Tomato spot wilt virus Tomato spotted wilt virus (TSWV) can infect more than 900 species of plants belonging to 70 families. The disease is distributed all over the world. RNA L is a negative RNAs, and its complementary chain encodes a RNA dependent RNA polymerase, RdRpSs, which is a double sense RNA. The complementary chain encodes nucleocapsid protein (RNA). These two diseases can cause serious damage to production and economic loss. Therefore, the molecular biological level of the two RNA viruses is further studied. The purpose of this study was to construct transgenic tobacco containing TSWV, RBSDV CP and RdRp proteins, and to establish its reverse genetic system. The pROK 鈪，

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