Ethylmethanesulfonate Saturation Mutagenesis in Arabidopsis to Determine Frequency of Herbicide Resistance

Abstract

Plant resistance to glyphosate has been reported far less frequently than resistance to sulfonylurea and imidazolinone herbicides. However, these studies tend to be anecdotal, without side by side comparisons for a single species or natural isolate. In this study, we tested the frequencies of resistance of three herbicides in a controlled ethylmethanesulfonate (EMS) saturation mutagenesis experiment, allowing a direct comparison of the frequencies at which resistant mutant plants arise. The 100% growth inhibition dose rates of glyphosate, chlorsulfuron (a sulfonylurea herbicide), and imazethapyr (an imidazolinone herbicide) were determined for Arabidopsis. Populations of EMS-mutagenized M 2 seedlings were sprayed with twice the 100% growth inhibition dose of glyphosate, chlorsulfuron, or imazethapyr, and herbicide-resistant mutants were identified. Although there were no glyphosate-resistant mutants among M 2 progeny of 125,000 Columbia and 125,000 Landsberg erecta M 1 lines, chlorsulfuron resistance and imazethapyr resistance each appeared at frequencies of 3.2 ϫ 10 Ϫ5 . Given the observed frequency of herbicide resistance mutations, we calculate that there are at least 700 mutations in each EMS-mutagenized Arabidopsis line and that fewer than 50,000 M 1 lines are needed to have a 95% chance of finding a mutation in any given G:C base pair in the genome. As part of this study, two previously unreported Arabidopsis mutations conferring resistance to imidazolinone herbicides, csr1-5 (Ala-122-Thr) and csr1-6 (Ala-205-Val), were discovered. Neither of these mutations caused enhanced resistance to chlorsulfuron in Arabidopsis.Spontaneous herbicide resistance is generally thought to occur within weed populations as a consequence of the intense selective pressure exerted by a lack of diversity in weed management practices (Gressel and Segel, 1978). Factors such as extended residual soil activity, lack of rotation to other herbicidal modes of action, and specific managerial practices further discriminate between resistant and susceptible individuals within a population (Powles and Holtum, 1994). In addition, the rate and severity at which resistant weed infestations occur is influenced by genetic and ecophysiological determinants such as the mode of inheritance of a given resistance mechanism, the absence or presence of fitness penalties associated with resistance, and the reproductive habit of a given weed species (Gressel and Segel, 1978;Jasieniuk et al., 1996; Gardner et al., 1998). To date, more than 261 herbicide-resistant weed biotypes exist distributed among 52 different countries, involving at least 17 different herbicide modes of action (Heap, 2002). Because application rate and other factors vary greatly in the field, it is difficult to make a direct comparison of the frequencies at which weeds develop resistance to different herbicides. To circumvent this problem, we have used a controlled laboratory setting to compare the frequencies at which heavily mutagenized populations of Arabidopsis develop resistanc...

High survival frequencies at low herbicide use rates in populations of Lolium rigidum result in rapid evolution of herbicide resistance

“…Additionally, a single cycle of selection with diclofop-methyl increased populationlevel resistance to a range of other herbicides (Figure 2), demonstrating the potential for rapid selection of diverse cross-resistance patterns. These observed phenotypic frequencies of resistance were orders of magnitude greater than rates of spontaneous target-site mutations, which give rise to target-site ALS resistance in Arabidopsis thaliana (Haughn and Somerville, 1987;Jander et al, 2003) and Nicotiana tabacum (Harms and DiMaio, 1991). These frequencies were also much greater than the initial frequency of target-site based resistance to ALS herbicides in three previously untreated L. rigidum populations (Preston and Powles, 2002).…”

Epistatic Interactions Among Herbicide Resistances in Arabidopsis thaliana: The Fitness Cost of Multiresistance

“…This cross design was replicated six times, giving six triple-heterozygous resistant-susceptible (R/S) plants (called the G2 family), each used to originate a segregating family (G3). The EMS origin of the csr1-1, ixr1-2, and axr1-3 lines means that they may carry several mutations other than the ones conferring resistance ( Jander et al 2003). By our crossing protocol, any EMS mutations other than the ones conferring resistance (except those closely linked to the resistance mutations) would contribute equally to the average fitness of each of the 27 genotypes.…”

The effects of the genetic background on herbicide resistance fitness cost and its associated dominance in Arabidopsis thaliana

“…To limit maternal effects among crosses, all F1 plants were grown at the same time in a greenhouse, under natural light supplemented by artificial light to provide a 16-h photoperiod and temperature maintained between 20 and 25 1C. The EMS origin of the axr1-3 line means that it may carry several mutations other than the ones conferring resistance (Jander et al, 2003). In a previous study, EMS mutations other than the one conferring axr1-3 resistance were found not to induce a significant reduction of plant fitness .…”