Chapter 26 Effects of Brassinolide on Conditioning and Germination of Witchweed (Striga asiatica) Seeds 1
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Y. Takeuchi , A. D. Worsham , and A. E. Awad Downloaded by UNIV OF ARIZONA on January 9, 2013 | http://pubs.acs.org Publication Date: November 4, 1991 | doi: 10.1021/bk-1991-0474.ch026
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Weed Control Research Institute, Utsunomiya University, Utsunomiya 321, Japan Crop Science Department, North Carolina State University, Raleigh, NC 27695-7620 International Institute of Tropical Agriculture, Ibadan 5320, Nigeria 2
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Brassinolide applied at an early stage of conditioning, shortened the conditioning period required before witchweed seeds would germinate after exposure to a germination stimulant. Although dl-strigol, natural stimulants from sorghum (Sorghum bicolor L. Moench) root exudate, kinetin, ethephon and gibberellin A applied after conditioning work as germination stimulants, they inhibit both conditioning and germination when applied during the conditioning period. Brassinolide eliminated the inhibitory effects on conditioning and subsequent germination caused by these chemicals applied during the conditioning period. The combination of brassinolide with either kinetin or ethephon induced higher germination of the seeds during the conditioning period than kinetin or ethephon alone. Brassinolide applied after conditioning in creased the rate of seed germination induced by the stimulants. Brassinolide eliminated the inhibitory effects of seed germination caused by indoleacetic acid and light. 3
Witchweed is a very troublesome root parasite of many warm-season grass crops in cluding corn (Zea mays L.), sorghum and sugarcane (Saccharum officinarum L.) (4). Reviews of seed germination of parasitic angiosperms have recently been published (33,39,40). In nature in soil, witchweed seeds will not germinate unless they are ex posed to natural stimulants releasedfromroots of host and many non-host plants (4, 15,17, 38-40). Sorgoleone and its analogues (5) from sorghum, a host species and d/-strigol (6, 7)fromcotton (Gossypium hirsutum L.), a non-host were isolated and their structures identified. Some analogues of dl-strigol, (75,23), cytokinins (75,57, 42), abscisic acid (ABA) (27), ethylene (10) and ethephon (9, 10) (an ethyleneproducing compound) were also found to induce the germination of witchweed seeds. Gibberellin A (GA ) induced the germination of scarified seeds, but did not induce the germination of non-scarified seeds while indoleacetic acid (IAA) inhibited seed germi nation (77). Witchweed seeds must be kept in a moist environment at 20 to 30°C for 1-3 weeks before the stimulants can initiate germination (4,15,39,40). This process is defined 3
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0097-6156/91/0474-0298$06.00/0 © 1991 American Chemical Society In Brassinosteroids; Cutler, H., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1991.
26. TAKEUCHI ET AL.
Conditioning and Germination of Witchweed Seeds
as conditioning by Hsiao et al. (15). It has been proposed that conditioning may (a) promote synthesis of a germination stimulant (3), (b) cause leaching of germination inhibitors from the seeds (2, 3, 8, 29, 35, 36), or (c) increase the permeability of a structure within the seeds, apparently the aleurone layer (35,39). After completing the conditioning process(es), seeds can respond to exposure to germination stimulants (4, 15-18, 39). Stimulants such as d/-strigol, cytokinins, ethylene, ethephon and G A , applied during the early stage of conditioning inhibited the conditioning process which in turn inhibited subsequent germination of the seed (15-18,20,21). ABA and IAA showed no apparent effect on the conditioning process (21). Zeatin induced 3 to 20% germination of seeds during conditioning. However germination of the remaining seeds was inhibited when exposed to the usual stimulants (27). A higher concentration of stimulants (75, 77,27,37) and a longer exposure time (75, 77,27) wererequiredto initiate germination if seeds were conditioned in the presence of certain stimulants compared to seeds conditioned in water (75, 77,27,37). The effects of many compounds have been tested on the conditioning of the seeds, but no promotive compound has beenreportedto date. Many studies have been conducted to determine the effectiveness of brassinosteroids as growthregulatorsin agricultural production systems (7, 72, 73, 30, 32, 41). However, the promotive effects of brassinosteroids on germination of weed have not been reported. This paper describes studies on the effects of brassinolide alone or in combination with various plant growthregulatorson conditioning and germination of witchweed seeds (34).
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Materials and Methods Seeds. Witchweed seeds were collected from plants in experimental field of the USDA Methods Development Laboratory, Whiteville, N.C., USA and stored for three years at room temperature. Viability of these seeds was more than 95% according to a tetrazolium chloride viability test. Chemicals. d/-Strigol was obtained in 1974fromDr. C. J. Sih of the University of Wisconsin and stored at 4°C. Brassinolide was obtainedfromDr. Ikegawa of Tokyo Institute of Technology in 1982 and stored at 4°C. Natural stimulants were obtained as follows: Five sorghum seeds ('Dekalb 64') were placed in sand in 1000 ml plastic cups. Each cup had five 1 mm-holes in the bottom and was placed in another cup of the same size. The seedlings were grown in a greenhouse and 200 ml of distilled water were added during the incubation. After 14 days, 15 ml of solution containing root exudatesfromsorghum had leaked into the outer cup. The solution was filtered and used as the natural stimulant. Seed Conditioning and Germination. Five hundred witchweed seeds were placed in a 9 cm-Petri dish and immersed in 5 ml of conditioning solution at 30°C for 3 to 15 days. After the conditioning period, the seeds were rinsed with distilled water and the wash water wasremoved.Groups of 50 seeds each were transferred to a 6-cm Petri dish containing a sheet of filter paper wetted with 2 ml of the terminal treatment solution. The dishes were placed in the dark at 30°C for 24 h. After the terminal treatment, germination of the seeds was determined under a lOx binocular microscope. Seeds were considered to have germinated if the radicle had emergedfromthe seed coat. Standard errors of the means were computed for all treatments which had three replications each and data were presented as the means of three separated experiments. In studying the effects if chemicals on germination, solution containing the chemicals were used as the terminal treatments for 24 h after seeds were conditioned in water for either 7 or 14 days. In studying the effects of chemicals on the conditioning of seeds and their subsequent germination, solutions containing the chemicals were used during conditioning and then stimulants (dl-strigol or natural stimulants) were applied as the terminal treatments. Percent germination wasrecordedafter 24 h.
In Brassinosteroids; Cutler, H., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1991.
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BRASSINOSTEROIDS: CHEMISTRY, BIOACTIVITY, AND APPLICATIONS
Results and Discussion Effects of Brassinolide and Combinations of Brassinolide with Various Plant Growth Regulators on Germination of the Seeds Conditioned in Water. Brassinolide at 10 to 10 M did not induce the germination of seeds conditioned in water for 7 or 14 days. However treatment with 10 M d/-strigol and undiluted natural stimulants (sorghum root exudate) resulted in approximately 60% germination of seeds conditioned in water for 7 days and more than 90% germination of seeds conditioned in water for 14 days (Table I). 15
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Table I. Germination of Witchweed Seeds Conditioned in Water at 30°C for 7 and 14 Days Followed by 24-h Terminal Treatment with Brassinolide, rf/-Strigol, and Natural Stimulants at 30°C in the Dark Terminal treatment Germination (%) Period of conditioning 14 days 7 days Water 0 6 Brassinolide 10-15M 0 0 Brassinolide 10 M 0 0 Brassinolide 10-»M 0 0 Brassinolide 10-^M 0 0 Brassinolide 10- M 0 0 Brassinolide 10-5M 0 0 d/-Strigol 63±4 94±3 10- M Natural stimulants diluted 4 x 85±5 28±2 90±4 Natural stimulants diluted 2 x 43±3 undiluted natural stimulants 63±4 94±3 13
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Two-and 4-fold-diluted natural stimulants, 1 0 M strigol, 6 x 10" and 2 x lO^M ethephon, and lO^M kinetin induced 28 to 43%, 54%, 61 and 30%, and 38% germination, respectively. When brassinolide was used with these plant growth regulators, it greatly increased seed germination, especially at 10* M (Table II). IAA used as a terminal treatment at 10 M did not induce germination. When used in combination 7
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Table II. Germination of Witchweed Seeds Conditioned in Water at 30°C for 7 Days Followed by 24-h Terminal Treatments with Combinations of Brassinolide (BR) with Natural Stimulants (STM), dlStrigol (ST), Ethephon (ET), Kinetin, or Gibberellin (GA ) at 30°C in the Dark Terminal treatment Germination (%) Terminal treatment Germination (%) Water 54±5 ST 10- M 0 80+3 BR 10-9M ST 10- M + BR 10-»M 0 BR lO-'M 83±6 ST 10-">M + BR lO-'M 0 30±4 STM diluted 4 x ET 2 x l ( H M 28±4 45±2 ET 2 x l ( H M + BR 10" M STMdiluted4x + BR 10- M 75±6 63±5 STMdiluted4x + BR 10- M 90±5 ET 2 x l ( H M + BR 10- M 61±2 STM diluted 2 x ET 6 X K H M 43±7 STMdiluted2x + BR l O ' M 80±4 ET 6xlCHM + BR 10" M 65±3 STMdiluted2x + BR 10 M 91±5 ET 6 x l ( H M + BR 10" M 84±5 38±3 GA 1(HM Kinetin 6x KHM 0 G A K H M + BR 10- M Kinetin 2 x l ( H M + BR 10" M 43±4 0 G A HHM + BR 10- M Kinetin 2 x l ( H M + BR 10 M 83±6 0 3
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In Brassinosteroids; Cutler, H., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1991.
26. TAKEUCHI ET AL.
Conditioning and Germination of Witchweed Seeds
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with 10" M d/-strigol or natural stimulants, IAA had inhibitory effects on germination induced by d/-strigol or natural stimulants. When used with d/-strigol and IAA or with natural stimulants and IAA, brassinolide eliminated the inhibitory effect of IAA on germination (Table III). These results indicated that brassinolide does not act as a stimulant for the germination of conditioned seeds, but that it enhances the effect of stimulants on germination. Brassinolide may increase the sensitivity of the seeds to stimulants by promoting some physiological processes, e.g. activation of endogenous growth substances (23). Table III. Germination of Witchweed Seeds Conditioned in Water at 30°C for 7 Days Followed by 24-h Terminal Treatments with Combinations of Brassinolide (BR) and Indoleacetic Acid (IAA) with
