Table_1_Characterization, Expression Profiling, and Functional Analysis of PtDef, a Defensin-Encoding Gene From Populus trichocarpa.XLSX

PtDef cloned from Populus trichocarpa contained eight cysteine domains specific to defensins. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) analysis showed that PtDef was expressed in all tissues tested, with lower expression in leaves and higher expression in petioles, stems, and roots. Purified fused PtDef inhibited Aspergillus niger, Alternaria Nees, Mucor corymbifer, Marssonina populi, Rhizopus sp., and Neurospora crassa. PtDef also inhibited the growth of Escherichia coli by triggering autolysis. PtDef overexpression in Nanlin895 poplar (Populus × euramericana cv. Nanlin895) enhanced the level of resistance to Septotinia populiperda. qRT-PCR analysis also showed that the expression of 13 genes related to salicylic acid (SA) and jasmonic acid (JA) signal transduction differed between transgenic and wild-type (WT) poplars before and after inoculation, and that PR1-1 (12–72 h), NPR1-2, TGA1, and MYC2-1 expression was higher in transgenic poplars than in WT. During the hypersensitivity response (HR), large amounts of H2O2 were produced by the poplar lines, particularly 12–24 h after inoculation; the rate and magnitude of the H2O2 concentration increase were greater in transgenic lines than in WT. Overall, our findings suggest that PtDef, a defensin-encoding gene of P. trichocarpa, could be used for genetic engineering of woody plants for enhanced disease resistance.

从毛果杨（Populus trichocarpa）克隆得到的PtDef含有8个防御素特有的半胱氨酸结构域。定量逆转录聚合酶链反应（qRT-PCR）分析显示，PtDef在所有检测组织中均有表达，叶片中表达量较低，而叶柄、茎秆及根系中表达量较高。纯化的融合PtDef能够抑制黑曲霉（Aspergillus niger）、链格孢属（Alternaria Nees）、伞状毛霉（Mucor corymbifer）、杨盘二孢菌（Marssonina populi）、根霉属（Rhizopus sp.）以及粗糙脉孢菌（Neurospora crassa）的生长。PtDef还可通过诱导自溶抑制大肠杆菌（Escherichia coli）的增殖。在南林895杨（Populus × euramericana cv. Nanlin895）中过表达PtDef，可提升其对杨生褐盘二孢菌（Septotinia populiperda）的抗病水平。qRT-PCR分析进一步显示，接种前后转基因与野生型（WT）杨树中，13个与水杨酸（SA）和茉莉酸（JA）信号转导相关的基因表达存在差异；且转基因杨树中PR1-1（12~72 h）、NPR1-2、TGA1以及MYC2-1的表达量高于野生型杨树。在超敏反应（HR）过程中，杨树株系会产生大量过氧化氢（H₂O₂），尤其在接种后12~24小时最为显著；转基因株系的H₂O₂浓度上升速率与幅度均大于野生型株系。综上，本研究结果表明，PtDef作为毛果杨的防御素编码基因，可用于木本植物的遗传工程改造以增强抗病性。