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A novel W1999S mutation and non-target site resistance impact on acetyl-CoA carboxylase inhibiting herbicides to varying degrees in a UK Lolium multiflorum population.

Kaundun SS, Bailly GC, Dale RP, Hutchings SJ, McIndoe E - PLoS ONE (2013)

Bottom Line: Additionally population UK21 is characterised by other resistance mechanisms, very likely non non-target site based, affecting several aryloxyphenoxyproprionate (FOP) herbicides but not the practical field rate of pinoxaden.The positive identification of wild type tryptophan and mutant serine alleles at ACCase position 1999 could be readily achieved with an original DNA based derived cleaved amplified polymorphic sequence (dCAPS) assay that uses the same PCR product but two different enzymes for positively identifying the wild type tryptophan and mutant serine alleles identified here.This paper highlights intrinsic differences between ACCase inhibiting herbicides that could be exploited for controlling ryegrass populations such as UK21 characterised by compound-specific target site and non-target site resistance.

View Article: PubMed Central - PubMed

Affiliation: Syngenta, Jealott's Hill International Research Centre, Bracknell, United Kingdom. deepak.kaundun@syngenta.com

ABSTRACT

Background: Acetyl-CoA carboxylase (ACCase) inhibiting herbicides are important products for the post-emergence control of grass weed species in small grain cereal crops. However, the appearance of resistance to ACCase herbicides over time has resulted in limited options for effective weed control of key species such as Lolium spp. In this study, we have used an integrated biological and molecular biology approach to investigate the mechanism of resistance to ACCase herbicides in a Lolium multiflorum Lam. from the UK (UK21).

Methodology/principal findings: The study revealed a novel tryptophan to serine mutation at ACCase codon position 1999 impacting on ACCase inhibiting herbicides to varying degrees. The W1999S mutation confers dominant resistance to pinoxaden and partially recessive resistance to cycloxydim and sethoxydim. On the other hand, plants containing the W1999S mutation were sensitive to clethodim and tepraloxydim. Additionally population UK21 is characterised by other resistance mechanisms, very likely non non-target site based, affecting several aryloxyphenoxyproprionate (FOP) herbicides but not the practical field rate of pinoxaden. The positive identification of wild type tryptophan and mutant serine alleles at ACCase position 1999 could be readily achieved with an original DNA based derived cleaved amplified polymorphic sequence (dCAPS) assay that uses the same PCR product but two different enzymes for positively identifying the wild type tryptophan and mutant serine alleles identified here.

Conclusion/significance: This paper highlights intrinsic differences between ACCase inhibiting herbicides that could be exploited for controlling ryegrass populations such as UK21 characterised by compound-specific target site and non-target site resistance.

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Related in: MedlinePlus

dCAPS procedures for the detection of the wild type tryptophan and mutant serine amino acid residues at ACCase codon position 1999: (a) XcmI restricted (134 bp) and unrestricted (164 bp) fragments correspond to sensitive W1999 and resistant S1999 ACCase alleles respectively.(b) MnlI restricted fragment is indicative of the mutant serine allele (120 bp) while the undigested band (164 bp) corresponds to the wild type tryptophan allele. Heterozygous plants display one each of the restricted and unrestricted PCR fragment in both assays.
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pone-0058012-g002: dCAPS procedures for the detection of the wild type tryptophan and mutant serine amino acid residues at ACCase codon position 1999: (a) XcmI restricted (134 bp) and unrestricted (164 bp) fragments correspond to sensitive W1999 and resistant S1999 ACCase alleles respectively.(b) MnlI restricted fragment is indicative of the mutant serine allele (120 bp) while the undigested band (164 bp) corresponds to the wild type tryptophan allele. Heterozygous plants display one each of the restricted and unrestricted PCR fragment in both assays.

Mentions: A dCAPS assay was developed for large scale genotyping of single UK21 plants in view of confirming the association between the 1999 mutation and pinoxaden resistance. Digestion with two different enzymes allowed the positive identification of the wild type tryptophan or mutant serine alleles. Typical dCAPS profiles as resolved on 2% agarose gel electrophoresis are provided in figures 2a and 2b. PCR generated a 164 bp fragment for all plants. When digested with the enzyme Xcm1, wild type homozygous plants showed a single restricted band of 134 bp (and 30 bp fragment not visible of 2% agarose gel) while plants that are homozygous for the serine 1999 allele displayed the undigested PCR fragment of 164 bp. As expected, heterozygous plants showed a copy each of the 164 and 134 bp fragments. The inverse restriction profiles were obtained with the enzyme MnlI with plants containing the wild type tryptophan allele being unrestricted whilst individuals with the mutant serine allele showed a shorter digested band of 120 bp. It is noteworthy that the difference in size of restricted PCR fragments between XcmI and MnlI treatments is due to the fact that the second enzyme generally cuts 6/7 bp downstream of the CCTC recognition site while the first enzyme cleaves 4/5 base pairs upstream of the TGG 1999 codon. The dCAPS and sequencing results for the 16 UK21 and 8 STD1 plants were totally correlated demonstrating the accuracy of the dCAPS assay. Genotyping of the 32 remaining UK21 plants (that were phenotyped with pinoxaden at a single recommended rate) with the XcmI based dCAPS assay confirmed the complete association between the presence of the mutated serine 1999 ACCase allele and resistance to pinoxaden (Table 1).


A novel W1999S mutation and non-target site resistance impact on acetyl-CoA carboxylase inhibiting herbicides to varying degrees in a UK Lolium multiflorum population.

Kaundun SS, Bailly GC, Dale RP, Hutchings SJ, McIndoe E - PLoS ONE (2013)

dCAPS procedures for the detection of the wild type tryptophan and mutant serine amino acid residues at ACCase codon position 1999: (a) XcmI restricted (134 bp) and unrestricted (164 bp) fragments correspond to sensitive W1999 and resistant S1999 ACCase alleles respectively.(b) MnlI restricted fragment is indicative of the mutant serine allele (120 bp) while the undigested band (164 bp) corresponds to the wild type tryptophan allele. Heterozygous plants display one each of the restricted and unrestricted PCR fragment in both assays.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585232&req=5

pone-0058012-g002: dCAPS procedures for the detection of the wild type tryptophan and mutant serine amino acid residues at ACCase codon position 1999: (a) XcmI restricted (134 bp) and unrestricted (164 bp) fragments correspond to sensitive W1999 and resistant S1999 ACCase alleles respectively.(b) MnlI restricted fragment is indicative of the mutant serine allele (120 bp) while the undigested band (164 bp) corresponds to the wild type tryptophan allele. Heterozygous plants display one each of the restricted and unrestricted PCR fragment in both assays.
Mentions: A dCAPS assay was developed for large scale genotyping of single UK21 plants in view of confirming the association between the 1999 mutation and pinoxaden resistance. Digestion with two different enzymes allowed the positive identification of the wild type tryptophan or mutant serine alleles. Typical dCAPS profiles as resolved on 2% agarose gel electrophoresis are provided in figures 2a and 2b. PCR generated a 164 bp fragment for all plants. When digested with the enzyme Xcm1, wild type homozygous plants showed a single restricted band of 134 bp (and 30 bp fragment not visible of 2% agarose gel) while plants that are homozygous for the serine 1999 allele displayed the undigested PCR fragment of 164 bp. As expected, heterozygous plants showed a copy each of the 164 and 134 bp fragments. The inverse restriction profiles were obtained with the enzyme MnlI with plants containing the wild type tryptophan allele being unrestricted whilst individuals with the mutant serine allele showed a shorter digested band of 120 bp. It is noteworthy that the difference in size of restricted PCR fragments between XcmI and MnlI treatments is due to the fact that the second enzyme generally cuts 6/7 bp downstream of the CCTC recognition site while the first enzyme cleaves 4/5 base pairs upstream of the TGG 1999 codon. The dCAPS and sequencing results for the 16 UK21 and 8 STD1 plants were totally correlated demonstrating the accuracy of the dCAPS assay. Genotyping of the 32 remaining UK21 plants (that were phenotyped with pinoxaden at a single recommended rate) with the XcmI based dCAPS assay confirmed the complete association between the presence of the mutated serine 1999 ACCase allele and resistance to pinoxaden (Table 1).

Bottom Line: Additionally population UK21 is characterised by other resistance mechanisms, very likely non non-target site based, affecting several aryloxyphenoxyproprionate (FOP) herbicides but not the practical field rate of pinoxaden.The positive identification of wild type tryptophan and mutant serine alleles at ACCase position 1999 could be readily achieved with an original DNA based derived cleaved amplified polymorphic sequence (dCAPS) assay that uses the same PCR product but two different enzymes for positively identifying the wild type tryptophan and mutant serine alleles identified here.This paper highlights intrinsic differences between ACCase inhibiting herbicides that could be exploited for controlling ryegrass populations such as UK21 characterised by compound-specific target site and non-target site resistance.

View Article: PubMed Central - PubMed

Affiliation: Syngenta, Jealott's Hill International Research Centre, Bracknell, United Kingdom. deepak.kaundun@syngenta.com

ABSTRACT

Background: Acetyl-CoA carboxylase (ACCase) inhibiting herbicides are important products for the post-emergence control of grass weed species in small grain cereal crops. However, the appearance of resistance to ACCase herbicides over time has resulted in limited options for effective weed control of key species such as Lolium spp. In this study, we have used an integrated biological and molecular biology approach to investigate the mechanism of resistance to ACCase herbicides in a Lolium multiflorum Lam. from the UK (UK21).

Methodology/principal findings: The study revealed a novel tryptophan to serine mutation at ACCase codon position 1999 impacting on ACCase inhibiting herbicides to varying degrees. The W1999S mutation confers dominant resistance to pinoxaden and partially recessive resistance to cycloxydim and sethoxydim. On the other hand, plants containing the W1999S mutation were sensitive to clethodim and tepraloxydim. Additionally population UK21 is characterised by other resistance mechanisms, very likely non non-target site based, affecting several aryloxyphenoxyproprionate (FOP) herbicides but not the practical field rate of pinoxaden. The positive identification of wild type tryptophan and mutant serine alleles at ACCase position 1999 could be readily achieved with an original DNA based derived cleaved amplified polymorphic sequence (dCAPS) assay that uses the same PCR product but two different enzymes for positively identifying the wild type tryptophan and mutant serine alleles identified here.

Conclusion/significance: This paper highlights intrinsic differences between ACCase inhibiting herbicides that could be exploited for controlling ryegrass populations such as UK21 characterised by compound-specific target site and non-target site resistance.

Show MeSH
Related in: MedlinePlus