RNA-Seq transcriptome analysis to identify genes involved in metabolism-based diclofop resistance in Lolium rigidum
RNA-Seq transcriptome analysis to identify genes involved in metabolism-based diclofop resistance in Lolium rigidum
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DOI：
10.1111/tpj.12514
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发表时间：
2014-06-01
期刊：
影响因子：
7.2
通讯作者：
Beffa, Roland
中科院分区：
文献类型：
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作者：
Gaines, Todd A.;Lorentz, Lothar;Beffa, Roland
除草剂抗性导致的杂草防除失败是一个日益严重的世界性问题，严重影响作物产量。杂草中基于代谢的除草剂抗性(称为代谢抗性)在遗传水平上还没有得到很好的表征。利用RNA-Seq转录组分析方法，在主要的全球杂草LoliumrigiumR的抗性种群(R)中寻找对除草剂敌敌畏产生代谢抗性的候选基因。组装了一个参考cDNA转录组(19623个重叠群)，并指定了推定的注释。使用来自未经处理的对照、仅使用佐剂的对照和三氯杀螨磷处理的R和易感(S)的Illumina Reads来测量全局基因表达。选择显示未经处理的R和未经处理的S的结构性表达差异的重叠群进行进一步的验证分析，包括11个假定被注释为细胞色素P450(细胞色素P450)、谷胱甘肽转移酶(GST)或葡萄糖基转移酶(GT)的重叠群，以及另外17个具有与代谢或信号转导相关的注释的重叠群。在正向遗传学验证实验中，从分离的F2群体中有9个重叠群在R个体中结构性上调，包括3个CytP450，1个硝酸单加氧酶(NMO)，3个GST和1个GT。利用这9个重叠群的主成分分析区分了F2-R和F2-S个体。在一个生理验证实验中，由于代谢增加，2，4-D预处理在S个体中诱导了双氯杀的保护作用，在9个遗传验证的重叠群中，有7个被显著诱导。四个重叠群(两个CytP450、NMO和GT)在9个田间进化的代谢抗性刚性乳杆菌种群中持续高表达。这四个重叠群与抗性表型密切相关，是导致代谢性双氯杀磷抗性的主要候选基因。
Weed control failures due to herbicide resistance are an increasing and worldwide problem that significantly affect crop yields. Metabolism-based herbicide resistance (referred to as metabolic resistance) in weeds is not well characterized at the genetic level. An RNA-Seq transcriptome analysis was used to find candidate genes that conferred metabolic resistance to the herbicide diclofop in a diclofop-resistant population (R) of the major global weed Loliumrigidum. A reference cDNA transcriptome (19623 contigs) was assembled and assigned putative annotations. Global gene expression was measured using Illumina reads from untreated control, adjuvant-only control, and diclofop treatment of R and susceptible (S). Contigs that showed constitutive expression differences between untreated R and untreated S were selected for further validation analysis, including 11 contigs putatively annotated as cytochrome P450 (CytP450), glutathione transferase (GST), or glucosyltransferase (GT), and 17 additional contigs with annotations related to metabolism or signal transduction. In a forward genetics validation experiment, nine contigs had constitutive up-regulation in R individuals from a segregating F2 population, including three CytP450, one nitronate monooxygenase (NMO), three GST, and one GT. Principal component analysis using these nine contigs differentiated F2-R from F2-S individuals. In a physiological validation experiment in which 2,4-D pre-treatment induced diclofop protection in S individuals due to increased metabolism, seven of the nine genetically validated contigs were induced significantly. Four contigs (two CytP450, NMO, and GT) were consistently highly expressed in nine field-evolved metabolic resistant L.rigidum populations. These four contigs were strongly associated with the resistance phenotype and are major candidates for contributing to metabolic diclofop resistance.