关键词: 双抗虫基因, 欧美杨107杨, 外源基因表达, Bt毒蛋白, 抗虫性

Abstract: [Objective] In order to identify and analyze the expression of exogenous genes in transgenic Populus×euramericana cv. '74/76' carrying bivalent insect-resistant genes BtCry1Ac and API (Abbreviation:Transgenic 107 poplar), and to select transgenic 107 poplar lines with high resistance to Lepidoptera pests Hlyphantria cunea and Clostera anachoreta.[Method]Two-year-old field seedlings of ten transgenic 107 poplar lines were selected as materials and non-transgenic 107 poplar as control. Detections of exogenous gene expression were conducted, including PCR detection, absolute fluorescence quantitative PCR, toxin detection and comparative studies on insect resistance of Lepidoptera pests.[Result]PCR detection showed that exogenous genes BtCry1Ac and API existed in the ten transgenic lines, but not in the CK, indicating that BtCry1Ac and API were steadily inserted into transgenic plant genome. The fluorescence quantitative PCR showed transcription abundance of BtCry1Ac in five lines PB1, PB2, PB6, PB9, and PB10 were higher, ranging from 1.72×10⁸ to 7.91×10⁹, PB1 was the highest; and followed by PB3, PB7, and PB8, ranging from 1.03×10⁷ to 2.35×10⁷; PB4, PB5, and CK detected no transcriptional expression of BtCry1Ac. ELISA toxin detection showed the content of Cry1Ac toxin expression was in accordance with variation tendency of transcription abundance. PB1 was the highest, with 665.11 ng·g^-1, PB4, PB5, and CK detected no toxin. The indoor insect feeding test showed that the insecticidal effect of transgenic 107 poplar on H. cunea larvae was higher than that of C. anachoreta. Eight transgenic lines expressed toxin had high resistance to L₁-L₆ (L₁-L₆ represent 1 to 6 instars)H. cunea larvae with a mortality of 100%, but there were differences on lethal time among these lines, the lethal time of PB1 and PB2 on every instars were shorter. PB1, PB2, PB6, PB9 and PB10 had a high resistance to L₁-L₄ C. anachoreta larvae with a mortality of 100%, the lethal time of PB1 and PB2 were shorter. The mortality of PB3, PB7, and PB8 on L₁-L₂C.anachoreta larvae was 100%, but the mortality of L₃-L₄ C. anachoreta larvae was as high as 22.22%. The 8 transgenic lines were divided into two resistance levels:PB1, PB2, PB6, PB9 and PB10 were high resistant lines, which showed high insect resistance to H. cunea and C. anachoreta; PB3, PB7, and PB8 were moderate resistant lines, which showed moderate insect resistance to C. anachoreta, but high resistance to H. cunea.[Conclusion] A series of molecular detections and indoor insect feeding tests were conducted on transgenic 107 poplar in this study. The results indicated that exogenous genes were steadily inserted into the poplar genome, and the 8 lines were of high-efficiency expression. Insect resistance test showed that transgenic 107 poplar had high resistance to H. cunea and C. anachoreta. Five lines were selected and showed high resistance to two pests, the other three lines PB3, PB7, and PB8 showed high resistance to H. cunea, but low or moderate resistance to C. anachoreta.

Key words: bivalent insect-resistant genes, Populus×euramericana cv. '74/76', exogenous gene expression, Bt toxin, insect resistance