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Identification of Reference Genes for Studying Herbicide Resistance Mechanisms in Japanese Foxtail (Alopecurus japonicus )

Published online by Cambridge University Press:  30 June 2017

Hongle Xu ,
Jun Li ,
Renhai Wu ,
Wangcang Su ,
Xibao Wu ,
Lingyue Wang  and
Liyao Dong
Hongle Xu
Affiliation:
Research Associate, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China, and Institute of Plant Protection, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China
Jun Li
Affiliation:
Associate Professor, Graduate Student, Graduate Student, and Professor, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China
Renhai Wu
Affiliation:
Associate Research Fellow and Research Associate, Institute of Plant Protection, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China
Wangcang Su
Affiliation:
Associate Research Fellow and Research Associate, Institute of Plant Protection, Henan Academy of Agricultural Sciences, Zhengzhou 450002, China
Xibao Wu
Affiliation:
Associate Professor, Graduate Student, Graduate Student, and Professor, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China
Lingyue Wang
Affiliation:
Associate Professor, Graduate Student, Graduate Student, and Professor, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China
Liyao Dong*
Affiliation:
Associate Professor, Graduate Student, Graduate Student, and Professor, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China
*
*Corresponding author’s E-mail: dly@njau.edu.cn
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Abstract

Herbicide-resistant weeds pose a considerable threat to agriculture, but their resistance mechanisms are poorly understood. Differential gene expression analysis of a weed subjected to herbicide treatment is a key step toward more mechanistic studies. Such an analysis, often involving quantitative real-time PCR (qPCR), requires suitable reference genes as internal controls. In this study, we identified optimal reference genes in the noxious weed, Japanese foxtail. This weed has evolved resistance to acetyl-coenzyme A carboxylase (ACCase) inhibitors. We analyzed the stability of eight commonly used candidate reference genes (glyceraldehyde-3-phosphate dehydrogenase [GAPDH]; ubiquitin [UBQ]; capsine phosphatase [CAP]; beta-tubulin [TUB]; eukaryotic initiation factor 4a [EIF4A]; elongation factor-1 alpha [EF1]; 18S ribosomal RNA [18S]; 25S ribosomal RNA [25S]) from root, stem, and leaf tissue of plants that were either resistant or sensitive to ACCase inhibitors, with or without herbicide stress, using qPCR. The results were further ranked and analyzed using geNorm, NormFinder, and BestKeeper software. These analyses identified EF1 and UBQ in roots, EF1, TUB, CAP, and 18S in stems, and EF1, GAPDH, and 18S in leaves as suitable references for qPCR normalization. We have identified a set of reference genes that can be used to study herbicide resistance mechanisms in Japanese foxtail.

Keywords

Japanese foxtailAlopecurus japonicus Steud.ACCaseBestKeeperfenoxaprop-P-ethylgene expressiongeNormNormFinderqPCR.

Information

Type
Physiology/Chemistry/Biochemistry
Information
Weed Science , Volume 65 , Issue 5 , September 2017 , pp. 557 - 566
Copyright
© Weed Science Society of America, 2017 

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Footnotes

Associate Editor for this paper: Patrick J. Tranel, University of Illinois.

References

Literature Cited

Andersen, CL, Jensen, JL, Orntoft, TF (2004) Normalization of real-time quantitative reverse transcription-PCR data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. Cancer Res 64:5245–5250 CrossRefGoogle Scholar
Bradley, KW, Wu, J, Hatzios, KK, Hagood, ES Jr (2001) The mechanism of resistance to aryloxyphenoxypropionate and cyclohexanedione herbicides in a johnsongrass biotype. Weed Sci 49:477–484 Google Scholar
Brunner, AMAB, Yakovlev, IAIY, Strauss, SHSS (2004) Validating internal controls for quantitative plant gene expression studies. BMC Plant Biol 4:14 Google Scholar
Bustin, SA (2002) Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems. J Mol Endocrinol 29:1 Google Scholar
Bustin, SA, Beaulieu, JF, Huggett, J, Jaggi, R, Kibenge, F, Olsvik, PA, Penning, LC, Toegel, S (2010) MIQE precis: practical implementation of minimum standard guidelines for fluorescence-based quantitative real-time PCR experiments. BMC Mol Biol 11:74 CrossRefGoogle ScholarPubMed
Bustin, SA, Benes, V, Garson, JA, Hellemans, J, Huggett, J, Kubista, M, Mueller, R, Nolan, T, Pfaffl, MW, Shipley, GL, Vandesompele, J, Wittwer, CT (2009) The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55:611–622 Google Scholar
Cha, TS, Najihah, MG, Bin Sahid, I, Chuah, TS (2014) Molecular basis for resistance to ACCase-inhibiting fluazifop in Eleusine indica from Malaysia. Pestic Biochem Phys 111:7–13 CrossRefGoogle ScholarPubMed
Czechowski, T, Stitt, M, Altmann, T, Udvardi, MK, Scheible, WR (2005) Genome-wide identification and testing of superior reference genes for transcript normalization in Arabidopsis . Plant Physiol 139:5–17 Google Scholar
Délye, C (2005) Weed resistance to acetyl coenzyme A carboxylase inhibitors: an update. Weed Sci 53:728–746 CrossRefGoogle Scholar
Délye, C, Jasieniuk, M, Le Corre, V (2013) Deciphering the evolution of herbicide resistance in weeds. Trends Genet 29:649–658 CrossRefGoogle ScholarPubMed
Dombrowski, JE, Martin, RC (2009) Evaluation of reference genes for quantitative RT-PCR in Lolium temulentum under abiotic stress. Plant Sci 176:390–396 Google Scholar
Duhoux, A, Délye, C (2013) Reference genes to study herbicide stress response in Lolium sp.: up-regulation of P450 genes in plants resistant to acetolactate-synthase inhibitors. PLoS One 8:e635765 Google Scholar
Fu, JX, Wang, Y, Huang, H, Zhang, C, Dai, SL (2013) Reference gene selection for RT-qPCR analysis of Chrysanthemum lavandulifolium during its flowering stages. Mol Breeding 31:205–215 CrossRefGoogle Scholar
Gaines, TA, Zhang, WL, Wang, DF, Bukun, B, Chisholm, ST, Shaner, DL, Nissen, SJ, Patzoldt, WL, Tranel, PJ, Culpepper, AS, Grey, TL, Webster, TM, Vencill, WK, Sammons, RD, Jiang, JM, Preston, C, Leach, JE, Westra, P (2010) Gene amplification confers glyphosate resistance in Amaranthus palmeri . Proc Natl Acad Sci USA 107:1029–1034 Google Scholar
Gutierrez, L, Mauriat, M, Guenin, S, Pelloux, J, Lefebvre, JF, Louvet, R, Rusterucci, C, Moritz, T, Guerineau, F, Bellini, C, Van Wuytswinkel, O (2008) The lack of a systematic validation of reference genes: a serious pitfall undervalued in reverse transcription-polymerase chain reaction (RT-PCR) analysis in plants. Plant Biotechnol J 6:609–618 CrossRefGoogle Scholar
Hong, S, Seo, PJ, Yang, M, Xiang, F, Park, C (2008) Exploring valid reference genes for gene expression studies in Brachypodium distachyon by real-time PCR. BMC Plant Biol 8:112 CrossRefGoogle ScholarPubMed
Jarosova, J, Kundu, JK (2010) Validation of reference genes as internal control for studying viral infections in cereals by quantitative real-time RT-PCR. BMC Plant Biol 10:146 CrossRefGoogle ScholarPubMed
Jiang, Q, Wang, F, Li, MY, Ma, J, Tan, GF, Xiong, AS (2014) Selection of suitable reference genes for qPCR normalization under abiotic stresses in Oenanthe javanica (BI.) DC. PLoS One 9:e922623 Google Scholar
Kong, QS, Yuan, JX, Niu, PH, Xie, JJ, Jiang, W, Huang, Y, Bie, ZL (2014) Screening suitable reference genes for normalization in reverse transcription quantitative real-time PCR analysis in melon. PLoS One 9:e871071 Google Scholar
Lee, JM, Roche, JR, Donaghy, DJ, Thrush, A, Sathish, P (2010) Validation of reference genes for quantitative RT-PCR studies of gene expression in perennial ryegrass (Lolium perenne L.). BMC Mol Biol 11:8 Google Scholar
Lin, YL, Lai, ZX (2010) Reference gene selection for qPCR analysis during somatic embryogenesis in longan tree. Plant Sci 178:359–365 Google Scholar
Livak, KJ, Schmittgen, TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCt method. Methods 25:402–408 CrossRefGoogle Scholar
Mohamed, IA, Li, RZ, You, ZG, Li, ZH (2012) Japanese foxtail (Alopecurus japonicus) resistance to fenoxaprop and pinoxaden in China. Weed Sci 60:167–171 Google Scholar
Nolan, T, Hands, RE, Bustin, SA (2006) Quantification of mRNA using real-time RT-PCR. Nature Protocols 1:1559–1582 Google Scholar
Ovesna, J, Kucera, L, Vaculova, K, Strymplova, K, Svobodova, I, Milella, L (2012) Validation of the β-amy1 transcription profiling assay and selection of reference genes suited for a RT-qPCR assay in developing barley caryopsis. PLoS One 7:e418867 Google Scholar
Petit, C, Pernin, F, Heydel, J, Délye, C (2012) Validation of a set of reference genes to study response to herbicide stress in grasses. BMC Res Notes 5:18 Google Scholar
Pfaffl, MW, Tichopad, A, Prgomet, C, Neuvians, TP (2004) Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: BestKeeper—Excel-based tool using pair-wise correlations. Biotechnol Lett 26:509–515 CrossRefGoogle ScholarPubMed
Powles, SB, Yu, Q (2010) Evolution in action: plants resistant to herbicides. Annu Rev Plant Biol 61:317–347 Google Scholar
Rechsteiner, MP, Widmer, F, Kolliker, R (2006) Expression profiling of Italian ryegrass (Lolium multiflorum Lam.) during infection with the bacterial wilt inducing pathogen Xanthomonas translucens pv. graminis . Plant Breeding 125:43–51 Google Scholar
Rozen, S, Skaletsky, H (2000) Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol 132:365–386 Google Scholar
Sato, K, Shin-I, T, Seki, M, Shinozaki, K, Yoshida, H, Takeda, K, Yamazaki, Y, Conte, M, Kohara, Y (2009) Development of 5006 full-length cDNAs in barley: a tool for accessing cereal genomics resources. DNA Res 16:81–89 Google Scholar
Schmittgen, TD, Livak, KJ (2008) Analyzing real-time PCR data by the comparative CT method. Nature Protocols 3:1101–1108 CrossRefGoogle Scholar
Silver, N, Best, S, Jiang, J, Thein, SL (2006) Selection of housekeeping genes for gene expression studies in human reticulocytes using real-time PCR. BMC Mol Biol 7:33 Google Scholar
Singh, R, Green, MR (1993) Sequence-specific binding of transfer-RNA by glyceraldehyde-3-phosphate dehydrogenase. Science 259:365–368 CrossRefGoogle ScholarPubMed
Soderlund, C, Descour, A, Kudrna, D, Bomhoff, M, Boyd, L, Currie, J, Angelova, A, Collura, K, Wissotski, M, Ashley, E, Morrow, D, Fernandes, J, Walbot, V, Yu, Y (2009) Sequencing, mapping, and analysis of 27,455 maize full-length cDNAs. PLoS Genetics 5:e100074011 Google Scholar
Tang, H, Li, J, Dong, L, Dong, A, Lü, B, Zhu, X (2012) Molecular bases for resistance to acetyl-coenzyme A carboxylase inhibitor in Japanese foxtail (Alopecurus japonicus). Pest Manag Sci 9:1241–1247 Google Scholar
Tong, ZG, Gao, ZH, Wang, F, Zhou, J, Zhang, Z (2009) Selection of reliable reference genes for gene expression studies in peach using real-time PCR. BMC Mol Biol 10:71 Google Scholar
Vandesompele, J, De Preter, K, Pattyn, F, Poppe, B, Van Roy, N, De Paepe, A, Speleman, F (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3:research0034Google Scholar
Wu, X, Zhang, T, Pan, L, Wang, L, Xu, H, Dong, L (2016) Germination requirements differ between fenoxaprop-P-ethyl resistant and susceptible Japanese foxtail (Alopecurus japonicus) biotypes. Weed Sci 64:653–663 Google Scholar
Xu, H, Li, J, Zhang, D, Cheng, Y, Jiang, Y, Dong, L (2014) Mutations at codon position 1999 of acetyl-CoA carboxylase confer resistance to ACCase-inhibiting herbicides in Japanese foxtail (Alopecurus japonicus). Pest Manag Sci 70:1894–1901 Google Scholar
Xu, H, Zhu, X, Wang, H, Li, J, Dong, L (2013) Mechanism of resistance to fenoxaprop in Japanese foxtail (Alopecurus japonicus) from China. Pestic Biochem Physiol 107:25–31 Google Scholar
Xu, YY, Zhu, XW, Gong, YQ, Xu, L, Wang, Y, Liu, LW (2012) Evaluation of reference genes for gene expression studies in radish (Raphanus sativus L.) using quantitative real-time PCR. Biochem Bioph Res Co 424:398–403 Google Scholar
Yang, CH, Dong, LY, Li, J, Moss, SR (2007) Identification of Japanese foxtail (Alopecurus japonicus) resistant to haloxyfop using three different assay techniques. Weed Sci 55:537–540 Google Scholar
Yang, Q, Yin, JJ, Li, G, Qi, LW, Yang, FY, Wang, RG, Li, GJ (2014) Reference gene selection for qRT-PCR in Caragana korshinskii Kom. under different stress conditions. Mol Biol Rep 41:2325–2334 CrossRefGoogle ScholarPubMed
Young, CA, Bryant, MK, Christensen, MJ, Tapper, BA, Bryan, GT, Scott, B (2005) Molecular cloning and genetic analysis of a symbiosis-expressed gene cluster for lolitrem biosynthesis from a mutualistic endophyte of perennial ryegrass. Mol Genet Genomics 274:13–29 Google Scholar
Yuan, JS, Tranel, PJ, Stewart, CN (2007) Non-target-site herbicide resistance: a family business. Trends Plant Sci 12:6–13 CrossRefGoogle ScholarPubMed
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