Pulp or Potential? Magneto-Priming as a Seed Priming Technique Has Little Effect on Seedling Performance but Results in Significant Alteration of trans-Zeatin and cis-Zeatin in Soybean (Glycine max)
Round 1
Reviewer 1 Report
In the submitted manuscript, Capperauld et al. evaluated the effect of magneto-priming soybean seeds on characteristics of seedlings. A static magnetic field had no significant effect on growth (radical emergence rate, seedling mass, hypocotyl length, etc.), physiological parameters (photosynthetic rate, transpiration rate, etc.), chlorophyl and isopentenyl adenosine riboside concentrations. Magneto-priming significantly increased only trans-zeatin and cis-zeatin concentrations. The results of this study contradict previous reports. Thus, the effectiveness of magnetic field treatment of plant seeds to improve growth/resilience is debatable. Research is needed to study the effects of magnetic fields on plants at the molecular level.
The effect of cytokinins on plant growth and development usually depends not on absolute concentrations, but on the ratio to auxins. Why did the authors not analyze the IAA content?
L.613-626. The production of reactive oxygen species was not analyzed in the manuscript, so this paragraph should be removed.
The Supplementary Materials should include the titles of the figures.
Author Response
In the submitted manuscript, Capperauld et al. evaluated the effect of magneto-priming soybean seeds on characteristics of seedlings. A static magnetic field had no significant effect on growth (radical emergence rate, seedling mass, hypocotyl length, etc.), physiological parameters (photosynthetic rate, transpiration rate, etc.), chlorophyl and isopentenyl adenosine riboside concentrations. Magneto-priming significantly increased only trans-zeatin and cis-zeatin concentrations. The results of this study contradict previous reports. Thus, the effectiveness of magnetic field treatment of plant seeds to improve growth/resilience is debatable. Research is needed to study the effects of magnetic fields on plants at the molecular level.
The effect of cytokinins on plant growth and development usually depends not on absolute concentrations, but on the ratio to auxins. Why did the authors not analyze the IAA content?
The authors would like to thank Reviewer #1 for their constructive feedback and encouragement. For clarity and convenience, all changes to the manuscript in response to the recommendations made herein have been highlighted in yellow in the revised manuscript.
While the authors do agree with Reviewer #1 regarding the role of auxins in plant development, we believe that many, if not most, of the effects of cytokinins (CK’s) work in concert with auxins, do not necessarily directly include auxin. In fact, examination of the role of CK’s in plant development has been made and reviewed, independent of consideration of auxins:
- Kieber, J. J. & Schaller, G. E. (2018). Cytokinin signalling in plant development. Development, 145(4).
- Liu, Y., Zhang, M., Meng, Z., Wang, B., & Chen, M. (2020). Research Progress on the Roles of Cytokinin in Plant Response to Stress. International Journal of Molecular Sciences, 21(18).
- Yu, Y., Li, Y., Yan, Z., & Duan, X. (2022). The Role of Cytokinins in Plants Under Salt Stress. J Plant Growth Regul. 41(1).
- Goggin, D. E., Emery, R. J. N., Kurepin, L. V., Powles, S. B. (2015). A potential role for endogenous microflora in dormancy release, cytokinin metabolism and the response to fluridone in Lolium rigidum. Annals of Botany. 115(2): 293–301.
The authors maintain that the discrete but thorough investigation of cytokinin fluctuations in response to MP with steady magnetic flux densities still presents a productive and worthwhile new body of evidence in an emerging field, even in the absence of auxin analysis.
L.613-626. The production of reactive oxygen species was not analyzed in the manuscript, so this paragraph should be removed.
The authors agree with the recommendations of Reviewer #1. L. 608-626 in the original manuscript have been removed.
The Supplementary Materials should include the titles of the figures.
The authors agree. This has been corrected.
Reviewer 2 Report
The manuscript by Capperauld et al. is devoted to verification of efficiency of magneto-priming of seeds of soybean. Authors did not reveal positive influence of this priming on plant growth, transpiration, and photosynthesis. This result is contradicted to other works showing efficiency of the magneto-priming and show, at least, necessity of following investigation of efficiency of the magneto-priming. It is very interesting; however, I have some comments and questions.
Main points:
1. My main doubt is related to results of photosynthetic measurements. Authors used light with intensity 2000 µmol m-2 s-1 to photosynthetic investigation (line 226); however, it is too intensive light. Based on the Farquhar—von Caemmerer—Berry, it can be expected that CO2 carboxylation by RuBisCo should limit photosynthetic processes; i.e. changes in photosynthetic light reactions should not influence CO2 assimilation in this case. On the other hand, photosynthetic light reactions can be primary target of magnetic field (e.g., through influence electron pair forming or through changes in ion transport across thylakoid membranes). It means that the influence of the magneto-priming on photosynthesis is not probable to be shown under used experimental conditions in case of changes in photosynthetic light reactions. Additionally, this light intensity should induce strong energy-dependent non-photochemical quenching for 3 min; induction of some photodamage is not fully excluded, too. Finally, it is not clear: Did this intensity of actinic light at photosynthetic measurements correspond intensity of light, which was used to cultivate soybean plants. Cultivation of soybean under 2000 µmol m-2 s-1 light intensity does not seem to be probable; however, if other light intensity was used for cultivation, why was this intensity not used to measure photosynthetic parameters? These points should be strongly clarified. I suppose that using light curves of CO2 assimilation, which can be analyzed on basis of the Farquhar—von Caemmerer—Berry, should be more informative; however, if only one light intensity was even used, why was it too intensive light?
2. I suppose that authors should explain using constant magnetic field in this work. Low-frequency magnetic fields are also used to modify plant growth and photosynthetic characteristics.
Specific points:
3. Section 2.4: What was light intensity used to cultivate soybean seedlings? What was the type of light source?
4. Section 2.5: What type of light was used as the actinic light (blue, red, white, or other)?
5. Figure 1c: What does the dotted line mean? Is it regression? It should be clarified.
Author Response
The manuscript by Capperauld et al. is devoted to verification of efficiency of magneto-priming of seeds of soybean. Authors did not reveal positive influence of this priming on plant growth, transpiration, and photosynthesis. This result is contradicted to other works showing efficiency of the magneto-priming and show, at least, necessity of following investigation of efficiency of the magneto-priming. It is very interesting; however, I have some comments and questions.
Main points:
- My main doubt is related to results of photosynthetic measurements. Authors used light with intensity 2000 µmol m-2 s-1 to photosynthetic investigation (line 226); however, it is too intensive light. Based on the Farquhar—von Caemmerer—Berry, it can be expected that CO2 carboxylation by RuBisCo should limit photosynthetic processes; i.e. changes in photosynthetic light reactions should not influence CO2 assimilation in this case. On the other hand, photosynthetic light reactions can be primary target of magnetic field (e.g., through influence electron pair forming or through changes in ion transport across thylakoid membranes). It means that the influence of the magneto-priming on photosynthesis is not probable to be shown under used experimental conditions in case of changes in photosynthetic light reactions. Additionally, this light intensity should induce strong energy-dependent non-photochemical quenching for 3 min; induction of some photodamage is not fully excluded, too. Finally, it is not clear: Did this intensity of actinic light at photosynthetic measurements correspond intensity of light, which was used to cultivate soybean plants. Cultivation of soybean under 2000 µmol m-2 s-1 light intensity does not seem to be probable; however, if other light intensity was used for cultivation, why was this intensity not used to measure photosynthetic parameters? These points should be strongly clarified. I suppose that using light curves of CO2 assimilation, which can be analyzed on basis of the Farquhar—von Caemmerer—Berry, should be more informative; however, if only one light intensity was even used, why was it too intensive light?
The authors would like to thank Reviewer #2 for their feedback. Responses to address each concern has been made separately below. For clarity and convenience, all changes to the manuscript in response to the recommendations made herein have been highlighted in yellow in the revised manuscript.
- I suppose that authors should explain using constant magnetic field in this work. Low-frequency magnetic fields are also used to modify plant growth and photosynthetic characteristics.
A section has been added to the introduction (L. 83-89) to clarify and support the use of steady magnetic field exposure, supported by relevant literature. Importantly, we believe that the results of research concerning magnetopriming (MP) of seeds with short intervals of steady magnetic flux and those which use prolonged treatments with low-frequency magnetic fields (LFMF) are not directly comparable. We believe that LFMF is an alternative crop enhancement method, requiring prolonged treatment in a controlled environment and therefore not immediately a field-deployable technique to implement at-scale due to technological (mainly energy) limitations. In the present work, the authors aimed to replicate the work of other investigators by studying the effects of MP as a seed priming method for enhancing germination and early vegetative growth.
Specific points:
Section 2.4: What was light intensity used to cultivate soybean seedlings? What was the type of light source?
Cultivation of soybean (ACC Mandor) was achieved using Sunblaster T5HO (54W) 6400K Light Reflector with NanoTech T5 Reflection System. The T5HO system is designed to provide full spectrum supplemental lighting, with enhanced spectral distribution relative intensity peaks at 435 nm and 615 nm. When measured using two independent calibrated light meters (Li-Cor, LI-250A), the photosynthetic photon flux density (PPFD) of full-spectrum light for our experiments was determined to be ~900 μmol/m²/s. These revisions have been made to section 2.4; L. 209-213.
Our previous inclusion of measurements of photosynthetically active radiation (PAR) has been removed as we did not directly assess PAR. Our previous measurement of 2,000 μmol/m²/s has also been updated to the true value (900 μmol/m²/s) throughout the manuscript. The previous value was obtained while the light intensity meter was programmed in the incorrect units for which it was not calibrated (lux).
Importantly, the intent of the authors was to examine the applicability of MP as a pre-sowing seed priming technique. Our investigation peered through the lens of assessing whether MP is readily field-deployable. We have intended to replicate the work of other investigators while using highly precise environmental and experimental controls including temperature and humidity, magnetic flux density, fixed exposure duration, number of seeds treated, and light intensity. We have set out to determine whether, under optimal environmental conditions (soil, humidity, temperature, and light), MP is decisively a growth-promoting pre-sow seed-priming technique. In the present work, we have provided a strong foundation upon which other groups may further elucidate the role of MP as a seed priming technique for growth under known and expected (intentional) environmental pressures which would preclude or be otherwise unfavourable for germination, including drought, salinity, or reduced supplemental lighting (100-500 μmol/m²/s).
Section 2.5: What type of light was used as the actinic light (blue, red, white, or other)?
Kindly refer to response above.
Figure 1c: What does the dotted line mean? Is it regression? It should be clarified.
The caption for Figure 1 has been updated to clarify the purpose of the dotted line in frame ‘c’.
Round 2
Reviewer 2 Report
Authors considered my comments. Particularly, 900 μmol m-2 s-1 light intensity seems to be possible for environmental conditions and to be suitable for photosynthetic measurements. I suppose that this interesting work can be accepted.
See "Major comments" to authors.
Author Response
"Authors considered my comments. Particularly, 900 μmol m-2 s-1 light intensity seems to be possible for environmental conditions and to be suitable for photosynthetic measurements. I suppose that this interesting work can be accepted."
The authors would like to thank Reviewer #2 for their constructive feedback, encouragement, and time. We are pleased with the decision of Reviewer #2.
