抗赤星病烟草的防卫基因的表达与基因组DNA结构的变化

Defence gene expression and DNA structure mutation in tobacco resistant to brown spot

用赤星病菌 (Alternariaalternata)毒性菌株产生的AT毒素诱导烟草 ,可使寄主获得对赤星病 80 .4 %的系统抗性 ,比病菌弱毒株TBA16孢子诱导的效果高约 2 0 %。烟草品种NC89心叶叶碟在含AT毒素 60 %和 75%的MS培养基上经 2轮胁迫筛选和植株再生 ,得到抗病品系NC89 TT。用 5种防卫基因的cDNA探针检测了不诱导、受AT毒素诱导、受病菌弱毒株激发子诱导的NC89和诱导与不诱导的NC89 TT第 4代植株 5种基因的转录活性 ,结果表明 ,苯丙氨酸氨裂解酶(PAL)基因在NC89中不能组成型表达 ,可被AT毒素诱导激活 ,但不受激发子诱导 ;病程相关蛋白(PR蛋白 )PR 1a、查尔酮合成酶 (CHS)、脂氧合酶 (LOX)基因在NC89中表现为诱导转录或诱导后转录活性增强 ;这 4种基因都能在NC89 TT中组成型表达。几丁质酶 (CHT)基因在NC89中组成型表达 ,但受诱导后及在NC89 TT中表达丧失。烟草基因组DNA的随机引物扩增 (RAPD)测定表明 ,AT毒素诱导了DNA结构的明显变化 ;与亲本NC89相比 ,NC89 TT的基因组DNA显示明显的序列重排或核苷酸替换。

AT toxin produced by a virulent strain TBA28 of tobacco brown spot pathogen Alternaria alternata induced systemic resistance being over 80% of a susceptible cultivar NC89. The resistance induced by the toxin was 20% higher than that by a hypovirulent strain TBA16 of the pathogen. A resistant somaclone NC89 TT had been produced by the process of tissue culture and plant regeneration, in which AT toxin was used as pressuring factor. NC89 TT was reproduced by seed production. Expressions of 5 defence genes were tested and DNA structural alternations were analyzed. Dot blots of RNAs from NC89 and NC89 TT plants either immunized by the elicitor and AT toxin or untreated were probed with cDNAs for pathogensis related (PR) protein PR 1a, chitinase (CHT), phenylalanine ammonia lyase (PAL), chalcone synthase (CHS), and lipoxygenase (LOX). Both PAL and PR la genes did not constitutively transcript in untreated NC89, and did in the fourth generation plants of NC89 TT. While, CHT gene was inverse. Transcription of PR la gene was activated by elicitor or AT toxin, and transcription of PAL gene only by the toxin .The genes of CHS and LOX showed constitutively feeble transcripts in untreated NC89, and were greatly enhanced in their transcript activity in NC89 plants induced by elicitor or AT toxin. A consistent tendency was found between the transcription activity and product accumulation of the genes indicated by PR la electrophoresis. Genome DNA structure mutation in different tobaccos was tested by random amplified polymorphic DNA (RAPD) using 5 primers. It was suggested that the sequence rearrangement or nucleotide substitution was responsible for the constitutive expression of the defence genes and such an expression resulted in resistance enhancement.