Materials and Reagents

1. Pipette tips
   
2. Plastic Petri dish (VWR, catalog number: 25384-326 )
   
3. Parafilm
   
4. 1.5 ml Eppendorf centrifuge tube (VWR, catalog number: 20170-355 )
   
5. Filter paper
   
6. Blue pestle (DWK Life Sciences, Kimble, catalog number: 749521-1500 )
   
7. Cuvette (VWR, catalog number: 414004-051 )
   
8. Syringe filters, 0.2 µm pore size (VWR, catalog number: 28145-475 )
   
9. Syringe
   
10. Arabidopsis thaliana ecotype Columbia (Col-0) and mutant line lhr1 (put3)
    
11. ScintiVerse^TM BD Cocktail (Fisher Scientific, catalog number: SX18-4 )
    
12. Clorox Bleach
    
13. Triton X-100 (Sigma-Aldrich, catalog number: T8787 )
    
14. (2,4-Dichlorophenoxy) acetic acid sodium salt monohydrate (Sigma-Aldrich, catalog number: D6679 )
    
15. Kinetin (Duchefa Biochemie, catalog number: K0905 )
    
16. Sodium hydroxide (NaOH)
    
17. Murashige & Skoog Basal Salt Mixture (PhytoTechnology Laboratories, catalog number: M524 )
    
18. Sucrose (Sigma-Aldrich, catalog number: S0389 )
    
19. Agar (Sigma-Aldrich, catalog number: A1296 )
    
20. Low melting point agarose (Sigma-Aldrich, catalog number: A9414 )
    
21. Methyl viologen dichloride hydrate (Sigma-Aldrich, catalog number: 856177 )
    
22. Spermidine (MP Biomedicals, catalog number: 02152068 )
    
23. Paraquat-methyl-¹⁴C dichloride hydrate (Sigma-Aldrich, catalog number: 313947 )
    Note: This product has been discontinued.
    
24. Spermidine trihydrochloride [Terminal Methylenes-3H (N)] (PerkinElmer, catalog number: NET522001MC )
    
25. Trizma^® base (Sigma-Aldrich, catalog number: T1503 )
    
26. Ethylenediaminetetraacetic acid disodium salt dihydrate (EDTA) (Sigma-Aldrich, catalog number: E5134 )
    
27. 2-Mercaptoethanol (Sigma-Aldrich, catalog number: M7522 )
    Note: This product has been discontinued.
    
28. Seed sterilization solution (see Recipes)
    
29. 2,000x 2,4-D (9.05 mM) (see Recipes)
    
30. 2,000x kinetin (1.86 mM) (see Recipes)
    
31. Callus induction solid medium (see Recipes)
    
32. Callus induction liquid medium (see Recipes)
    
33. ½ MS solid medium (see Recipes)
    
34. ½ MS liquid medium (see Recipes)
    
35. 200 µM non-¹⁴C-labeled PQ solution (see Recipes)
    
36. 2.04 mM non-³H-labeled Spd solution (see Recipes)
    
37. 40.37 µM non-³H-labeled Spd solution (see Recipes)
    
38. Solution 1: ¹⁴C-labeled PQ solution (see Recipes)
    
39. Solution 2: ¹⁴C-labeled PQ solution (see Recipes)
    
40. Solution 3: ³H-labeled Spd solution (see Recipes)
    
41. Solution 4: ³H-labeled Spd solution (see Recipes)
    
42. Solution 5: ³H-labeled Spd solution (see Recipes)
    
43. Solution 6: ³H-labeled Spd solution (see Recipes)
    
44. 1 M Tris-HCl, pH 7.5 (see Recipes)
    
45. 0.5 M EDTA, pH 8.0 (see Recipes)
    
46. Crude protein extraction buffer (see Recipes)
    

Equipment

1. Pipettes
   
2. pH meter
   
3. Weighing balance
   
4. Laminar flow hood
   
5. Stirring bar
   
6. Magnetic stirrer (VWR, model: 200 Mini-stirrer )
   
7. Vortex (Fisher Scientific, model: Vortex-Genie 2 )
   
8. Scintillation counter (Beckman Coulter, model: LS-6500 )
   
9. Centrifuge (Beckman Coulter, model: Microfuge^® 22R , catalog number: 368831)
   
10. Spectrophotometer (Bio-Rad Laboratories, model: SmartSpec Plus, catalog number: 1702525 )
    Note: This product has been discontinued.
    
11. Autoclave
    

Procedure