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Genetic Screens for Floral Mutants in Arabidopsis thaliana: Enhancers and Suppressors

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Flower Development

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1110))

Abstract

The flower is a hallmark feature that has contributed to the evolutionary success of land plants. Diverse mutagenic agents have been employed as a tool to genetically perturb flower development and identify genes involved in floral patterning and morphogenesis. Since the initial studies to identify genes governing processes such as floral organ specification, mutagenesis in sensitized backgrounds has been used to isolate enhancers and suppressors to further probe the molecular basis of floral development. Here, we first describe two commonly employed methods for mutagenesis (using ethyl methanesulfonate (EMS) or T-DNAs as mutagens), and then describe three methods for identifying a mutation that leads to phenotypic alterations—traditional map-based cloning, TAIL-PCR, and deep sequencing in the plant model Arabidopsis thaliana.

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References

  1. Laibach F (1943) Arabidopsis thaliana (L.) Heynh als objekt fur genetische und entwicklungsphysiologische Untersuchungen. Bot Archiv 44:439–455

    Google Scholar 

  2. Leutwiler LS, Hough-Evans BR, Meyerowitz EM (1984) The DNA of Arabidopsis thaliana. Mol Gen Genet 194:15–23

    Article  CAS  Google Scholar 

  3. Meyerowitz EM (1989) Arabidopsis, a useful weed. Cell 56(2):263–269

    Article  CAS  PubMed  Google Scholar 

  4. Meyerowitz EM, Pruitt RE (1985) Arabidopsis thaliana and plant molecular genetics. Science 229(4719):1214–1218

    Article  CAS  PubMed  Google Scholar 

  5. Pruitt RE, Meyerowitz EM (1986) Charac-terization of the genome of Arabidopsis thaliana. J Mol Biol 187(2):169–183

    Article  CAS  PubMed  Google Scholar 

  6. Bowman JL, Meyerowitz EM (1991) Genetic control of pattern formation during flower development in Arabidopsis. Symp Soc Exp Biol 45:89–115

    CAS  PubMed  Google Scholar 

  7. Bowman JL, Smyth DR, Meyerowitz EM (1989) Genes directing flower development in Arabidopsis. Plant Cell 1(1):37–52

    CAS  PubMed Central  PubMed  Google Scholar 

  8. Coen ES, Meyerowitz EM (1991) The war of the whorls: genetic interactions controlling flower development. Nature 353(6339):31–37

    Article  CAS  PubMed  Google Scholar 

  9. Meyerowitz EM et al (1991) A genetic and molecular model for flower development in Arabidopsis thaliana. Dev Suppl 1:157–167

    CAS  PubMed  Google Scholar 

  10. Drews GN, Weigel D, Meyerowitz EM (1991) Floral patterning. Curr Opin Genet Dev 1(2):174–178

    Article  CAS  PubMed  Google Scholar 

  11. Jack T, Brockman LL, Meyerowitz EM (1992) The homeotic gene APETALA3 of Arabidopsis thaliana encodes a MADS box and is expressed in petals and stamens. Cell 68(4):683–697

    Article  CAS  PubMed  Google Scholar 

  12. Smyth DR, Bowman JL, Meyerowitz EM (1990) Early flower development in Arabidopsis. Plant Cell 2(8):755–767

    CAS  PubMed Central  PubMed  Google Scholar 

  13. Weigel D et al (1992) LEAFY controls floral meristem identity in Arabidopsis. Cell 69(5):843–859

    Article  CAS  PubMed  Google Scholar 

  14. Yanofsky MF et al (1990) The protein encoded by the Arabidopsis homeotic gene agamous resembles transcription factors. Nature 346(6279):35–39

    Article  CAS  PubMed  Google Scholar 

  15. Bowman JL, Smyth DR (1999) CRABS CLAW, a gene that regulates carpel and nectary development in Arabidopsis, encodes a novel protein with zinc finger and helix-loop-helix domains. Development 126(11):2387–2396

    CAS  PubMed  Google Scholar 

  16. Sieburth LE, Running MP, Meyerowitz EM (1995) Genetic separation of third and fourth whorl functions of AGAMOUS. Plant Cell 7(8):1249–1258

    CAS  PubMed Central  PubMed  Google Scholar 

  17. Chen X, Meyerowitz EM (1999) HUA1 and HUA2 are two members of the floral homeotic AGAMOUS pathway. Mol Cell 3(3):349–360

    Article  CAS  PubMed  Google Scholar 

  18. Cheng Y et al (2003) Two RNA binding proteins, HEN4 and HUA1, act in the processing of AGAMOUS pre-mRNA in Arabidopsis thaliana. Dev Cell 4(1):53–66

    Article  CAS  PubMed  Google Scholar 

  19. Li J, Jia D, Chen X (2001) HUA1, a regulator of stamen and carpel identities in Arabidopsis, codes for a nuclear RNA binding protein. Plant Cell 13(10):2269–2281

    CAS  PubMed Central  PubMed  Google Scholar 

  20. Ji L et al (2011) ARGONAUTE10 and ARGONAUTE1 regulate the termination of floral stem cells through two microRNAs in Arabidopsis. PLoS Genet 7(3):e1001358

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  21. Chen X (2004) A microRNA as a translational repressor of APETALA2 in Arabidopsis flower development. Science 303(5666):2022–2025

    Article  CAS  PubMed  Google Scholar 

  22. Liu X et al (2011) AGAMOUS terminates floral stem cell maintenance in Arabidopsis by directly repressing WUSCHEL through recruitment of Polycomb Group proteins. Plant Cell 23(10):3654–3670

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  23. Siegfried KR et al (1999) Members of the YABBY gene family specify abaxial cell fate in Arabidopsis. Development 126(18):4117–4128

    CAS  PubMed  Google Scholar 

  24. Sawa S et al (1999) FILAMENTOUS FLOWER, a meristem and organ identity gene of Arabidopsis, encodes a protein with a zinc finger and HMG-related domains. Genes Dev 13(9):1079–1088

    Article  CAS  PubMed  Google Scholar 

  25. Eshed Y, Baum SF, Bowman JL (1999) Distinct mechanisms promote polarity establishment in carpels of Arabidopsis. Cell 99(2):199–209

    Article  CAS  PubMed  Google Scholar 

  26. Telfer A, Poethig RS (1998) HASTY: a gene that regulates the timing of shoot maturation in Arabidopsis thaliana. Development 125(10):1889–1898

    CAS  PubMed  Google Scholar 

  27. Wagner D, Meyerowitz EM (2002) SPLAYED, a novel SWI/SNF ATPase homolog, controls reproductive development in Arabidopsis. Curr Biol 12(2):85–94

    Article  CAS  PubMed  Google Scholar 

  28. Eshed Y et al (2001) Establishment of polarity in lateral organs of plants. Curr Biol 11(16):1251–1260

    Article  CAS  PubMed  Google Scholar 

  29. Muller R, Bleckmann A, Simon R (2008) The receptor kinase CORYNE of Arabidopsis transmits the stem cell-limiting signal CLAVATA3 independently of CLAVATA1. Plant Cell 20(4):934–946

    Article  PubMed Central  PubMed  Google Scholar 

  30. Brand U et al (2000) Dependence of stem cell fate in Arabidopsis on a feedback loop regulated by CLV3 activity. Science 289(5479):617–619

    Article  CAS  PubMed  Google Scholar 

  31. Nimchuk ZL, Tarr PT, Meyerowitz EM (2011) An evolutionarily conserved pseudokinase mediates stem cell production in plants. Plant Cell 23(3):851–854

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. Nag A, Yang Y, Jack T (2007) DORNRO-SCHEN-LIKE, an AP2 gene, is necessary for stamen emergence in Arabidopsis. Plant Mol Biol 65(3):219–232

    Article  CAS  PubMed  Google Scholar 

  33. Levin JZ et al (1998) A genetic screen for modifiers of UFO meristem activity identifies three novel FUSED FLORAL ORGANS genes required for early flower development in Arabidopsis. Genetics 149(2):579–595

    CAS  PubMed  Google Scholar 

  34. Page DR, Grossniklaus U (2002) The art and design of genetic screens: Arabidopsis thaliana. Nat Rev Genet 3(2):124–136

    Article  CAS  PubMed  Google Scholar 

  35. Papdi C et al (2010) Genetic screens to identify plant stress genes. Methods Mol Biol 639:121–139

    Article  CAS  PubMed  Google Scholar 

  36. Kim Y, Schumaker KS, Zhu JK (2006) EMS mutagenesis of Arabidopsis. Methods Mol Biol 323:101–103

    CAS  PubMed  Google Scholar 

  37. Krieg DR (1963) Ethyl methanesulfonate-induced reversion of bacteriophage T4rII mutants. Genetics 48:561–580

    CAS  PubMed  Google Scholar 

  38. Greene EA et al (2003) Spectrum of chemically induced mutations from a large-scale reverse-genetic screen in Arabidopsis. Genetics 164(2):731–740

    CAS  PubMed  Google Scholar 

  39. McCallum CM et al (2000) Targeted screening for induced mutations. Nat Biotechnol 18(4):455–457

    Article  CAS  PubMed  Google Scholar 

  40. Jander G et al (2003) Ethylmethanesulfonate saturation mutagenesis in Arabidopsis to determine frequency of herbicide resistance. Plant Physiol 131(1):139–146

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  41. Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16(6):735–743

    Article  CAS  PubMed  Google Scholar 

  42. Tzfira T et al (2004) Agrobacterium T-DNA integration: molecules and models. Trends Genet 20(8):375–383

    Article  CAS  PubMed  Google Scholar 

  43. Szabados L et al (2002) Distribution of 1000 sequenced T-DNA tags in the Arabidopsis genome. Plant J 32(2):233–242

    Article  CAS  PubMed  Google Scholar 

  44. Wang Y (2008) How effective is T-DNA insertional mutagenesis in Arabidopsis? J Biochem Tech 1(1):11–20

    CAS  Google Scholar 

  45. Li Y et al (2006) Analysis of T-DNA insertion site distribution patterns in Arabidopsis thaliana reveals special features of genes without insertions. Genomics 87(5):645–652

    Article  CAS  PubMed  Google Scholar 

  46. Weigel D et al (2000) Activation tagging in Arabidopsis. Plant Physiol 122(4):1003–1013

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  47. Radhamony RN, Prasad AM, Srinivasan R (2005) T-DNA insertional mutagenesis in Arabidopsis: a tool for functional genomics. Electr J Biotech 8(1). doi: 10.2225/vol8-issue1-fulltext-4

    Google Scholar 

  48. Lukowitz W, Gillmor CS, Scheible WR (2000) Positional cloning in Arabidopsis. Why it feels good to have a genome initiative working for you. Plant Physiol 123(3):795–805

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  49. Liu YG, Whittier RF (1995) Thermal asymmetric interlaced PCR: automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking. Genomics 25(3):674–681

    Article  CAS  PubMed  Google Scholar 

  50. Liu YG, Chen Y (2007) High-efficiency thermal asymmetric interlaced PCR for amplification of unknown flanking sequences. Biotechniques 43(5):649–654

    Article  CAS  PubMed  Google Scholar 

  51. Austin RS et al (2011) Next-generation mapping of Arabidopsis genes. Plant J 67(4):715–725

    Article  CAS  PubMed  Google Scholar 

  52. Schneeberger K et al (2009) SHOREmap: simultaneous mapping and mutation identification by deep sequencing. Nat Methods 6(8):550–551

    Article  CAS  PubMed  Google Scholar 

  53. Zhu Y et al (2012) Gene discovery using mutagen-induced polymorphisms and deep sequencing: application to plant disease resistance. Genetics 192(1):139–146

    Article  CAS  PubMed  Google Scholar 

  54. Sarin S et al (2008) Caenorhabditis elegans mutant allele identification by whole-genome sequencing. Nat Methods 5(10):865–867

    Article  CAS  PubMed Central  PubMed  Google Scholar 

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Author information

Authors and Affiliations

  1. Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, CA, USA

    Thanh Theresa Dinh, Elizabeth Luscher, Shaofang Li, Xigang Liu, So Youn Won & Xuemei Chen

  2. NSF ChemGen IGERT program, University of California, Riverside, CA, USA

    Thanh Theresa Dinh

  3. Howard Hughes Medical Institute, University of California, Riverside, CA, USA

    Xuemei Chen

Authors
  1. Thanh Theresa Dinh
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  2. Elizabeth Luscher
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  3. Shaofang Li
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  4. Xigang Liu
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  5. So Youn Won
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  6. Xuemei Chen
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Corresponding author

Correspondence to Xuemei Chen .

Editor information

Editors and Affiliations

  1. Center for Research in Agricultural Genomics, Barcelona, Spain

    José Luis Riechmann

  2. Trinity College Dublin, Ireland

    Frank Wellmer

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Dinh, T.T., Luscher, E., Li, S., Liu, X., Won, S.Y., Chen, X. (2014). Genetic Screens for Floral Mutants in Arabidopsis thaliana: Enhancers and Suppressors. In: Riechmann, J., Wellmer, F. (eds) Flower Development. Methods in Molecular Biology, vol 1110. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4614-9408-9_6

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  • DOI: https://doi.org/10.1007/978-1-4614-9408-9_6

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4614-9407-2

  • Online ISBN: 978-1-4614-9408-9

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