Establishing a herbicide-metabolizing enzyme library inBeckmannia syzigachneto identify genes associated with metabolic resistance

Abstract

Non-target site resistance (NTSR) to herbicides is an increasing concern for weed control. Metabolic herbicide resistance is an important mechanism for NTSR. However, little is known about metabolic resistance at the genetic level. In this study, we have identified three fenoxaprop-P-ethyl-resistant American sloughgrass (Beckmannia syzigachne Steud.) populations, in which the molecular basis for NTSR remains unclear. To reveal the mechanisms of metabolic resistance, the genes likely to be involved in herbicide metabolism (e.g. for cytochrome P450s, esterases, hydrolases, oxidases, peroxidases, glutathione S-transferases, glycosyltransferases, and transporter proteins) were isolated using transcriptome sequencing, in combination with RT-PCR (reverse transcription-PCR) and RACE (rapid amplification of cDNA ends). Consequently, we established a herbicide-metabolizing enzyme library containing at least 332 genes, and each of these genes was cloned and the sequence and the expression level compared between the fenoxaprop-P-ethyl-resistant and susceptible populations. Fifteen metabolic enzyme genes were found to be possibly involved in fenoxaprop-P-ethyl resistance. In addition, we found five metabolizing enzyme genes that have a different gene sequence in plants of susceptible versus resistant B. syzigachne populations. These genes may be major candidates for herbicide metabolic resistance. This established metabolic enzyme library represents an important step forward towards a better understanding of herbicide metabolism and metabolic resistance in this and possibly other closely related weed species. This new information may help to understand weed metabolic resistance and to develop novel strategies of weed management.

miR397/Laccase Gene Mediated Network Improves Tolerance to Fenoxaprop-P-ethyl in Beckmannia syzigachne and Oryza sativa

“…Consequently, many B. syzigachne populations have evolved resistance to fenoxaprop- P -ethyl, thereby rendering them as hard-to-control weeds (Pan et al, 2015b). In our previous studies, three fenoxaprop- P -ethyl-resistant B. syzigachne populations involving both TSR and NTSR were identified (Pan et al, 2015b), and our RNA-seq analysis detected structural changes and alterations in the transcriptional regulation of several genes encoding metabolizing enzymes (Pan et al, 2016). The RNA-seq analysis also showed that laccase gene expression was down-regulated in the three fenoxaprop- P -ethyl-resistant B. syzigachne populations.…”

“…Three fenoxaprop- P -ethyl-resistant B. syzigachne populations possessing both NTSR and TSR and one susceptible population (S) were used in this study (Pan et al, 2016). These populations were collected from the following locations: S from Feidong in Anhui Province, where fenoxaprop- P -ethyl was never applied; R1 from Xinlucun in Jiangsu Province with 10 years of fenoxaprop- P -ethyl use history; R2 from Lijiacun in Jiangsu Province, with 6 years of fenoxaprop- P -ethyl use; and R3 from Guocun in Jiangsu Province, with 5 years of fenoxaprop- P -ethyl use.…”

“…The resistance tillers were self-pollinated in isolation to produce seeds of progeny populations. Thus, three resistance-enriched populations (R1, R2, and R3) were generated (Pan et al, 2016). With this method, three “purified” progeny R populations were obtained (thus minimizing genetic variation unrelated to fenoxaprop- P -ethyl resistance).…”

“…Fenoxaprop- P -ethyl was applied at one-eighth of the recommended dose to allow survival of susceptible plants for gene expression analysis (Pan et al, 2016). The expression of candidate genes was measured in 30 untreated and 30 fenoxaprop- P -ethyl-treated plants for each fenoxaprop- P -ethyl dose at 24, 48, and 72 h. Total RNA was extracted using TRIzol (Invitrogen Inc., Carlsbad, CA, United States) following the manufacturer’s protocol.…”

“…NTSR can be mediated via several mechanisms (Délye, 2013; Yu and Powles, 2014), and often under polygenic control (Busi et al, 2013; Gaines et al, 2014; Duhoux et al, 2015; Gardin et al, 2015). Nevertheless, most recent studies have focused on metabolic resistance (Gaines et al, 2014; Duhoux et al, 2015; Gardin et al, 2015; Pan et al, 2016), and very few of them have investigated the molecular mechanisms underlying NTSR.…”