title: A high-throughput seed germination assay for root parasitic plants.
abstract: BACKGROUND
  Some root-parasitic plants belonging to the Orobanche, Phelipanche or Striga genus represent one of the most destructive and intractable weed problems to agricultural production in both developed and developing countries. Compared with most of the other weeds, parasitic weeds are difficult to control by conventional methods because of their life style. The main difficulties that currently limit the development of successful control methods are the ability of the parasite to produce a tremendous number of tiny seeds that may remain viable in the soil for more than 15 years. Seed germination requires induction by stimulants present in root exudates of host plants. Researches performed on these minute seeds are until now tedious and time-consuming because germination rate is usually evaluated in Petri-dish by counting germinated seeds under a binocular microscope.
  
  
  RESULTS
  We developed an easy and fast method for germination rate determination based on a standardized 96-well plate test coupled with spectrophotometric reading of tetrazolium salt (MTT) reduction. We adapted the Mosmann's protocol for cell cultures to germinating seeds and determined the conditions of seed stimulation and germination, MTT staining and formazan salt solubilization required to obtain a linear relationship between absorbance and germination rate. Dose-response analyses were presented as applications of interest for assessing half maximal effective or inhibitory concentrations of germination stimulants (strigolactones) or inhibitors (ABA), respectively, using four parameter logistic curves.
  
  
  CONCLUSIONS
  The developed MTT system is simple and accurate. It yields reproducible results for germination bioassays of parasitic plant seeds. This method is adapted to high-throughput screenings of allelochemicals (stimulants, inhibitors) or biological extracts on parasitic plant seed germination, and strengthens the investigations of distinctive features of parasitic plant germination.
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Jean-Bernard
authors.initials: JB
authors.lastname: Pouvreau
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Zachary
authors.initials: Z
authors.lastname: Gaudin
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Bathilde
authors.initials: B
authors.lastname: Auger
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Marc-Marie
authors.initials: MM
authors.lastname: Lechat
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Gauthier
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Delavault
authors.affiliation: Laboratoire de Biologie et de Pathologie Végétales EA 1157, SFR 4207 QUASAV, Nantes University, 44322 Nantes, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Simier
references.citation: J Agric Food Chem. 2010 Mar 10;58(5):2902-7
references.pmid: 20155908
references.citation: New Phytol. 2009 Jun;182(4):965-974
references.pmid: 19323792
references.citation: Pest Manag Sci. 2009 May;65(5):478-91
references.pmid: 19222046
references.citation: Plant Physiol. 2012 Aug;159(4):1524-44
references.pmid: 22723084
references.citation: J Exp Bot. 2012 Sep;63(14):5311-22
references.pmid: 22859674
references.citation: Ann Bot. 2005 Nov;96(6):1121-7
references.pmid: 16157629
references.citation: Bioorg Med Chem. 2011 Aug 15;19(16):5006-11
references.pmid: 21757362
references.citation: J Agric Food Chem. 2011 Sep 14;59(17):9226-31
references.pmid: 21819156
references.citation: J Agric Food Chem. 2009 Jul 8;57(13):5853-64
references.pmid: 19507857
references.citation: Mol Plant. 2013 Jan;6(1):38-62
references.pmid: 23204499
references.citation: Phytochemistry. 2007 Oct;68(19):2487-92
references.pmid: 17604061
references.citation: Phytopathology. 2003 Apr;93(4):451-7
references.pmid: 18944360
references.citation: Plant Cell Physiol. 2010 Jul;51(7):1095-103
references.pmid: 20403809
references.citation: J Immunol Methods. 1983 Dec 16;65(1-2):55-63
references.pmid: 6606682
references.citation: Pest Manag Sci. 2009 May;65(5):467-70
references.pmid: 19222028
references.citation: Cancer Res. 1988 Sep 1;48(17):4827-33
references.pmid: 3409223
references.citation: Phytochemistry. 2011 May;72(7):624-34
references.pmid: 21353686
references.citation: Pest Manag Sci. 2009 May;65(5):453-9
references.pmid: 19206075
references.citation: Pest Manag Sci. 2011 Aug;67(8):1015-22
references.pmid: 21480462
references.citation: Plant Sci. 2001 Apr;160(5):951-959
references.pmid: 11297792
references.citation: J Agric Food Chem. 2002 Mar 27;50(7):1911-7
references.pmid: 11902932
references.citation: Mol Plant Microbe Interact. 2012 Jul;25(7):993-1004
references.pmid: 22414435
references.citation: J Exp Bot. 2001 Nov;52(364):2227-34
references.pmid: 11604462
references.citation: Mol Plant. 2013 Jan;6(1):141-52
references.pmid: 23220943
references.citation: J Agric Food Chem. 2005 Mar 9;53(5):1598-603
references.pmid: 15740046