E-Book Overview
This collection of readily reproducible Arabidopsis protocols has been updated to reflect recent advances in plant biology, the completion of the Arabidopsis genome sequence, which is essential for studying plant function, and the development of whole systems approaches that allow global analysis of gene expression and protein and metabolite dynamics. The authors have included nearly all techniques developed in Arabidopsis, others recently adapted from the traditional work in crop species, and the most recent ones using Arabidopsis as a model system. Highlights include the most recent methods-transcriptomics, proteomics, and metabolomics - and their novel applications (phosphoproteomics, DNA microarray-based genotyping, high throughput metabolite profiling, and single-cell RNA).
E-Book Content
1 Growth of Plants and Preservation of Seeds Randy Scholl, Luz Rivero-Lepinckas,
and Deborah Crist
1. Introduction Arabidopsis can be grown in a variety of environmental settings including growth rooms, wmdow ledges, outdoors, growth chambers, and greenhouses. The growth media that can be employed include soil, commercial greenhouse mixes, vermrcuhte, and other relatively inert media watered wtth nutrient solutions, as well as agar. This chapter focuses on growth of plants on so11 in various envrromnental settings and especrally m growth chambers and greenhouses. Harvestmg, seed quaky, and seed preservation are also considered. Whereas Arubidopszs IS adaptable, it 1s not the equivalent of a house plant in that a number of environmental factors must be carefully controlled, or the culture may result m very unhealthy plants and even total failure. One of the main objectives of this chapter 1s to indrcate which conditions are critical to healthy plant growth and development and to high quality and quanttty of seeds. The plant and seed management methods are discussed m the chronological order m which they would normally be utilized.
2. Materials
2.1. Growth and Uarvesf 1. 2 3 4. 5 6 7 8.
Plastic pots (e.g , IO-cm square) (Hummert, Earth City, MO). Sol1 mrxture (e.g , Metromix 350 or other peat moss-based potting mix). Large autoclavable contamer. Large spoon or trowel pH Meter Magnetic stirring devrce 2M KOH or 1M H,SO, Beakers (100 or 250 mL, 1 L) From Methods m Molecular Brology, Vol 82 Arabrdopsrs Protocols Edrted by J Martinez-Zapater and J Sallnas 0 Humana Press Inc , Totowa.
1
NJ
2
Scholl, Rivero-Lepmckas,
and Cnst
9 Two magnetic stnrmg bars 10. Drstdled water. 11 Mtcronutrtent stock (used in making nutrient solution) To 90 mL of stirring, drstdled water add the following ingredients m order (concentrations shown are final concentration after volume has been adjusted). After the last salt has been added and is dissolved, adjust the volume to 100 mL. Ingredients 70 mM H,BO,, 14 mM MnCl,, 0.5 mA4 CuS04, 1 mA4ZnSO,, 0.2 mA4Na2Mo04, 10 mMNaC1, 0.01 mA4 CoCl,. 12. Nutrient: Add approx 700 mL of distilled water and a magnetic stnrmg bar to a 1-L beaker and stir Add salts m the amounts (final concentration basis) and order
shown: 5 mA4KN03,
13. 14. 15. 16
2.5 mA4KH2P0,
(adjusted to pH 6.5), 2.0 mMMgSO,,
2.0
nuI4 Ca(NO,),, 50 pJ4 Fe-EDTA, 1 mL micronutrient stock (step 11). After all salts have drssolved, adjust to pH 6 5 with 2MKOH (or lMH,SO,) and bring the volume to 1 L Solutron may be stored for several weeks before use, but should be discarded rf cloudmess or precipitates appear 8-cm round watch glass Pasteur prpet and latex bulb. Labeling tape. Permanent marker
2.2. Postharvest
Seed Management
1. Light-weight transparent plastic food storage bags (approx 4 L) 2. Hand sieves (mesh size = 0.425 mm). 3. Small glass jars (125 mL) or other storage containers (preferabl