Determination of the Phloem Path and Destination of the Photosynthates by Autoradiography and Fluorescent Marker Imaging

doi:10.21203/rs.3.rs-4304310/v1

Download PDF

Research Article

Determination of the Phloem Path and Destination of the Photosynthates by Autoradiography and Fluorescent Marker Imaging

https://doi.org/10.21203/rs.3.rs-4304310/v1

This work is licensed under a CC BY 4.0 License

Version 1

posted

You are reading this latest preprint version

Vascular bundle connectivity, which is closely associated with phyllotaxis, plays a crucial role in determine the destination of photosynthates. The correlation between vascular connections and phosphate distribution has been studied in some plants; however, it is far from fully understood. In this study, the phloem paths and destinations of photosynthates in soybean plants was investigated. In soybeans, trifoliate leaves are alternately arranged. To trace the movement of photosynthates, radioisotope-labeled 14C-sucrose was applied to mature leaves, and autoradiography was performed. The results indicated that photosynthates were translocated from the first trifoliate (L1) or second trifoliate (L2) to the immature third trifoliate (L3) and fourth trifoliate (L4) leaves, which are located on both side of the stem. When 14C -sucrose was added to one side of the source leaves, the 14C signal was detected on the corresponding side of the sink leaves. The photosynthates were translocated from the left side of L1 to the right side of L2 and the left side of L3. The phloem paths were traced using carboxyfluorescein (CF), which is known a phloem marker, and 14C-sucrose. The photosynthates were translocated via different phloem paths from both L1 and L2, as well as from the left and right side of the trifoliate. These results indicate that photosynthates from a single trifoliate are broadly distribute, while the underlying translocation paths are finely subdivided.

autoradiography

soybean

photosynthate

translocation

phyllotaxis

The translocation and partitioning of photosynthates are recognized as important mechanisms for plant growth. Photosynthesis occurs in leaves, with mature leaves producing more photosynthates than they consume. Photosynthates are exported from mature leaves (the source) and translocated via the phloem to sinks, which require these nutrients. The efficient translocation of photosynthates to sinks is directly associated with improved growth and yield.

The distribution of photosynthates is influenced by the sink and source locations, with vascular connections playing a significant role in their distribution(Shishido & Hori, 1977). Phyllotaxis, the arrangement pattern of leaves on a stem, is closely related to these vascular connections and determines the destination of photosynthates. A relationship between phyllotaxis and uneven distributions of photosynthates has been reported in model plants such as Arabidopsis(Kölling et al., 2013) and tobacco(Jones et al., 1959). In tomato, detailed studies have been reported on phyllotaxis, vascular connections, and their relationship to photosynthate translocation to the fruit(Shishido et al., 1988; Shishido & Hori, 1991). Similarly, in sour cherry, phyllotaxis and its influence on photosynthate translocation have been reported, although the specifics of transport to the fruit remain unclear (Kappes & Flore, 1989). The translocation of photosynthates from specific source leaves to fruit has been explored in melon (Barzegar et al., 2013) and apple (Sha et al., 2021), but the vascular connections involved have not been thoroughly investigated. Despite the great attention given to the importance of photosynthate translocation to fruit, this process in far from fully understood.

The soybean (Glycine max, Fabaceae family), an annual dicotyledonous plant, is widely consumed due to its high oil and protein content. As soybean is a significant source of plant protein and oil, increasing soybean yield is desirable(Hartman et al., 2011; Liu et al., 2020). Recent studies have focused on how sunlight, temperature, and carbon dioxide concentration affect soybean photosynthetic ability, growth, and yield(Burroughs et al., 2023; Drag et al., 2020; Feng et al., 2019; Hitz et al., 2019; Landau et al., 2022; Singh et al., 2021).

In the initial stages of development, the soybean plant forms a pair of cotyledons, succeeded by a pair of primary leaves positioned at right angles to the cotyledon. Subsequently, trifoliate leaflets, each with three leaflets, develop in sequence, alternating and positioned perpendicularly to the primary leaf (PL), each opening at an angle of 180°(Yoshikawa, 2019). Each leaf position n, n+2 and n+4 has the same orientation relative to the stem, which is directly opposite to the orientation of leaves at positions n+1, n+3, and n+5. However, due to the twisted nature of the stems and the significant changes in leaf arrangement caused by the pulvinus resulting in increased sunlight absorption, it is difficult to discern phyllotaxis in mature plants. Although numerous studies were performed from the 1950s to the 1980s to trace the ¹⁴C movement from the absorption of ¹⁴CO₂into soybean leaves(O. Biddulph & Cory, 1965; Fisher, 1970; Kagawa & Wong, 1985; Thaine et al., 1959; Thrower, 1962; Vernon & Aronoff, 1952), these studies focused on the distribution of the photosynthates in the whole plant, the timing of sink-to-source transition, or the types of carbon assimilated compounds. The relationships between source and sink locations and phyllotaxy were not mentioned. A comprehensive understanding of the translocation paths and destinations of photosynthates in soybean plants can enhance growth efficiency and increase yields.

In this study, we aimed to elucidate the translocation paths and destinations of photosynthates in soybean plants. Our study focused on soybean plants in the early stages of nutritional growth, a phase in which distinct phyllotaxis is identified. To elucidate the photosynthate destinations, ¹⁴C-sucrose was added to the source leaves, and ¹⁴C activity was measured. Furthermore, the distributions of ¹⁴C and CF in stems derived from different sources were compared to determine the individual path of photosynthates.

Plant material and growth conditions

Seeds of soybean (Glycine max. cv. Kosuzu) (Sato Masayuki Seed Company) were germinated in vermiculite and transferred to 1/2 Hoagland hydroponic solution. The plants were grown under a 16 h light/8 h dark cycle (280 μmol m-2 s -1) at 27 ˚C. The hydroponic solution was refreshed every week, and the plants were hydroponically cultivated for 8-14 days. To maintain experimental consistency, plant samples were selected based on their growth stage rather than the number of days of culture. The degree of expansion of the leaves was used as an indicator. When investigating photosynthate destinations, we focused on young leaves. Although L2 was expanding in both cases, L3 folding and unfolding states were distinguished. Thus, we expressed “L2-expanding” and “L3-unfolding”. When investigating the translocation pathway, we focused on the source leaves. More than 80 % of the expanded leaves were selected as the source. If L2 was > 80 % expanded and L3 was still expanding, it was described as “L2 > 80 % expanded”, regardless of the state of L4.

Application of ¹⁴CO₂

To trace the destination of the photosynthates, ¹⁴CO₂ was applied to L1. To prevent photosynthesis in parts other than L1, the remaining aboveground parts were covered with aluminum foil. The soybean plants were placed in an acrylic box connected to a vial through a tube. ¹⁴CO₂ gas was generated by reacting 5 MBq NaH¹⁴CO₃ (37 MBq/ml, 135 μl) (NEC086H005MC, PerkinElmer) and an excess of lactic acid (0.1 M, 150 μl) in the vial. The plants were exposed to ¹⁴CO₂ for 15 minutes. Subsequently, ¹⁴CO₂ was absorbed using NaOH. Afterward, the plants were moved from the acrylic box, and the aluminum foil was removed. Cultivation was continued for 24 hours under the growth conditions, after which the ¹⁴C activity in the plants was detected.

Application of ¹⁴C-sucrose and 5’ (6)-carboxyfluorescein diacetate (CFDA)

The selected leaves were gently abraded with sandpaper, followed by the application of ¹⁴C-sucrose (7.4 MBq/ml, 5 μl) (MC266, Moravek) and/or CFDA (1 mM, 5 μl) (19582, CAY). Subsequently, the leaf was covered with polyethylene film to prevent desiccation. The plants were cultivated under growth conditions for 4 to 6 hours to visualize the ¹⁴C signal in the stem and for 24 hours to detect the ¹⁴C activity within the aboveground part.

Imaging ¹⁴C and CF in plants

The distribution of the ¹⁴C signal in the aboveground part was detected by autoradiography. The plants were covered with polyethylene film and exposed to the imaging plate within a cassette at 4 ˚C for 3 hours to 3 days depending on the ¹⁴C activity. The ¹⁴C activity image was obtained by a fluorescent image analyzer (Amersham Typhoon, Cytiva). For stem imaging, freehand cross-sections were prepared using a razor blade. These sections were wrapped in polyethylene film, and CF images and bright field images were captured using a fluorescence microscope (BX60, Olympus) prior to ¹⁴C detection. Images of the ¹⁴C activity of the stems, as well as the aboveground parts, were obtained after the plants were exposed to imaging plates at -80 ˚C for 9 to 14 days. To identify the structure, cross-sections were stained with 0.05 % toluidine blue and observed by microscopy. These images were analyzed using ImageJ (version 1.53t). Imaging analyses were performed on more than three individual soybeans, and comparable results were obtained. Representative results are shown in each figure.

Distribution of ¹⁴C added as ¹⁴CO₂ gas and ¹⁴C-sucrose droplets

To ascertain whether carbon compounds derived from exogenously added sucrose behave the same as carbon assimilates produced within plants, we compared the distributions of ¹⁴C derived from ¹⁴CO₂ gas and ¹⁴C-sucrose droplets. ¹⁴CO₂ or ¹⁴C-sucrose was added to L1 when the plants were in the L3-expanding stage (Fig. 1). The results showed that the ¹⁴C signal was evenly distributed throughout the mesophyll of L1, which absorbed ¹⁴CO₂ (Fig. 1a). In contrast, L1, to which ¹⁴C-sucrose was applied, exhibited extremely high ¹⁴C activity at the application site, with less activity detected at the midvein and no activity detected at other site (Fig. 1b). ¹⁴C activity was detected in the petiole of L1, the main stem, the apex, L3 and the L3 petiole regardless of the chemical form of ¹⁴C. ¹⁴C activity was not detected in L2 or in the PL or cotyledons located below L1 in either case. The ¹⁴C distribution in plants treated with ¹⁴C-sucrose mirrored that of plants that absorbed ¹⁴CO₂, except L1, where the leaf received the ¹⁴C application. Subsequent ¹⁴C-tracing experiments were performed using only ¹⁴C-sucrose.

Translocation of photosynthates from source leaves at different leaf positions

To examine the destination of photosynthates from source leaves at different leaf positions, ¹⁴C-sucrose was added to L1 or L2 of the plants at the L4-unfolding stage (Fig. 2a, b). High ¹⁴C activity at L4 and moderate activity at L3 were detected in both plants with ¹⁴C-sucrose added to L1 and L2. Little ¹⁴C activity was detected in the cotyledons and PL in both cases. When ¹⁴C-sucrose was added to L1, L2 and the L2 petiole exhibited negligible ¹⁴C activity. When ¹⁴C-sucrose was added to L2, L1 and the L1 petiole showed minimal ¹⁴C activity. These results showed that the photosynthates from L1 and L2 were delivered to the same sink sites according to their sink activity; L4 had greater sink activity than L3. Moreover, L2, L1, PL and cotyledons acted as sources and did not receive photosynthates from other leaves.

Destination of photosynthates from different areas within a single trifoliate

The soybean trifoliate comprises one terminal leaflet and two lateral leaflets. In our ¹⁴C-sucrose experiment, we noted an uneven distribution of ¹⁴C between the left and right sides of the sink leaf when ¹⁴C-sucrose was applied only to one lateral leaflet. To determine whether this uneven left-right ¹⁴C distribution in the leaves was incidental of consistent, we applied ¹⁴C-sucrose to one lateral leaflet or to either side of the midrib within the terminal leaflet of L1 of the plants at the L3-unfolding stage (Fig. 2c). When ¹⁴C-sucrose was applied to the left side of the lateral leaflet, high ¹⁴C activity was observed on the right side of L2 and the left side of L3. Conversely, when ¹⁴C-sucrose was applied to the right side of the lateral leaflet, high ¹⁴C activity was observed on the left side of L2 and the right side of L3. When ¹⁴C-sucrose was added to each side of the terminal leaflet, L3 showed an uneven distribution of ¹⁴C-activity, similar to that observed when ¹⁴C-sucrose was added to the lateral leaflet. When ¹⁴C-sucrose was applied to all three leaflets of L1, ¹⁴C activity was detected throughout L2 and L3. These results showed that the destination of ¹⁴C depends on where ¹⁴C-sucrose was applied to the left or right side of the source leaf.

Movement of ¹⁴C-sucrose and CF in the stem

Photosynthates can be translocated from different sources to the same sink (Fig. 2a, b), and the destination area within a sink leaf differs depending on where photosynthates are loaded from within a source leaf (Fig. 2c). To determine the translocation paths of photosynthates from different sources, we performed imaging analysis using ¹⁴C-sucrose and CFDA (Fig. 3). CFDA, a cell-permeable and nonfluorescent compound, is cleaved by an intracellular esterase to form a cell-impermeable fluorescent substance, CF, which moves between cells only by plasmodesmata(Jiang et al., 2019). Since the distribution of foliar-absorbed CF is restricted to the phloem region(Grignon et al., 1989), it is commonly used as a phloem sap tracer. A mixture of ¹⁴C-sucrose and CFDA was applied to L1 of the plants at the L3-expanding stage. Cross sections of the petiole and main stem were prepared to observe the ¹⁴C and CF distributions (Fig. 3a). In the petiole, both ¹⁴C and CF signals were detected throughout the cambium (Fig. 3b). To support the understanding of signal distributions, each image in the main stem section was set with the L1 petiole connecting to the left, and the signal around the cambium was captured in a 360˚ clockwise direction starting from the 12:00 position (Fig. 3c). The signal intensities of ¹⁴C and CF are expressed as relative values ranging from 0 to 1. Signal differences represent the CF intensity minus the ¹⁴C intensity, ranging from -1 to 1. The signal differences will be close to 0 throughout if the distributions of the ¹⁴C and CF intensities match. At the L1 to L2 internodes, the ¹⁴C and CF intensities displayed three large peaks (Fig. 3c-iii). In the stem above the L3 and PL to L1, internode, the ¹⁴C and CF intensities did not exhibit three peaks, but the values fluctuated (Fig. 3c-ii, iv). In the stem above L3, near the apex, CF signals were detected only in the phloem, whereas ¹⁴C signals were also found outside the vascular bundle (Fig. 3c-ii). Comparison of ¹⁴C autoradiography and toluidine blue staining images revealed ¹⁴C in the xylem and epidermis in addition to the phloem (Fig. 3d, e). The graph shapes of the ¹⁴C and CF intensities showed similar patterns in all sections, indicating that ¹⁴C and CF passed through the same regions of the vascular bundles.

Translocation paths of photosynthates from different areas within a single source leaf

The destination area within a sink leaf differs depending on where photosynthates were loaded from within a source leaf (Fig. 2c). This is expected to be due to the distinct paths from different loading areas. To verify this, ¹⁴C-sucrose and CFDA were applied to the left and right lateral leaflets of L1 of the same plant at the L3-unfolding stage, and imaging analysis was subsequently conducted, as shown in Fig. 4a. The distributions of ¹⁴C and CF at the petioles and pulvinus were distinct (Fig. 4b). In the stem, both the ¹⁴C and CF intensities showed two peaks in the same region of the L1 to L2 internode (Fig. 4a-iii) and in the PL to L1 internode (Fig. 4a-iv). Both the ¹⁴C and CF intensities were greater in the 270 ˚ region, but the peak top positions did not overlap (Fig. 4c, red arrow). These results indicated that photosynthates unloaded from different areas in a source leaf were delivered to different areas in the same sink via different paths.

Translocation paths of photosynthates from leaves at different leaf positions

Photosynthates were observed to be translocated to various destinations via distinct pathways (Fig. 2c, Fig. 4). It was postulated that if the destination was the same, the translocation paths would also be the same. To validate this hypothesis, ¹⁴C-sucrose and CFDA were added to L1 and L2 in plants at the L2 >80 % expanded stage. Imaging analysis was subsequently conducted at each position (Fig. 5a). At the internodes of PL to L1 (Fig. 5a-iii) and L1 to L2 (Fig. 5a-ii), both ¹⁴C and CF intensities peaked in the same regions, showing three peaks, that did not overlap (Fig. 5b-ii and iii). However, the ¹⁴C and CF intensities showed two peaks at the L2 to L3, internodes (Fig. 5a-i), and the peak regions did not overlap with each other (Fig. 5b-i). These results suggest that photosynthates translocated from different sources pass through different regions of vascular bundles in the stem, even if the translocation sinks are the same. These findings were confirmed with similar experiments; the leaf positions of ¹⁴C-sucrose and CFDA were switched (Fig. S1).

Although the destinations of photosynthates from L1 and L2 in soybean were the same, the translocation paths were different (Figs. 2a, 5 and S1). L1 and L2 are located on opposite sides of the stem, which may account for the distinct vascular regions utilized within the stem. To verify this, we compared the translocation paths from L1 and L3, which are located on the same side of the stem. Imaging of stem sections was performed by foliar addition of ¹⁴C-sucrose and CFDA to L1 and L3, respectively, at the L3 > 80 % expanded stage. However, ¹⁴C and CF signals could not be clearly detected in the same sections (data not shown). This may be because the distance between L1 and L3 is long and the CF signal strength decreases with distance, so we could not find a place where we can detect the signal loaded from both L1 and L3. Therefore, we divided the experiment into two experiments, one comparing the translocation pathways from L1 and L2 and the other from L2 and L3 (Fig. S2a). Different plants at the same growth stage were used. For both plants, CFDA was added to L2, while ¹⁴C-sucrose was added to L1 or L3. In each experiment, clear signals of both ¹⁴C and CF were detected only in the internode between the two leaves with added CFDA and ¹⁴C-sucrose, respectively. In both images, the signals of ¹⁴C and CF intensities along the cambium showed three peaks, each of which did not overlap (Fig. S2b). These results suggested that photosynthates from L1 and L3 were translocated through vascular bundles in the same orthostichy.

Photosynthates synthesized in mature source leaves, are translocated to nutrient-demanding sinks for growth and maturation. Enhancing the distribution rate of these sinks can potentially improve growth and crop yield. However, the distribution rate of photosynthates is influenced by various factors, including the physiological state and location of the source and sink. It is known that the photosynthates from a particular source are preferentially translocated to sinks in the same orthostichy of the vascular system corresponding to phyllotaxis, resulting in their uneven distribution in the plant(Jones et al., 1959; Kappes & Flore, 1989; Kölling et al., 2013). However, only a limited number of species have been studied in detail regarding the destinations and translocation paths of photosynthates from specific sources. Applying Watson and Casper's hypothetical pathways of assimilate movement, and from several experimental results, photosynthate translocation in soybean is inferred to occur on the same side of the source leaves (Watson and Casper 1984) (see below) (Fig. 6a). Biddulph et al. reported that ³²P introduced into the L1 of soybean moved in the phloem of vascular bundles of the stem that were associated with the L3 leaf (S. Biddulph et al., 1958). In reproductive soybean, photosynthates from specific leaves are reportedly distributed to pods on the same node and every other node(Alan B. Bennett et al., 1984; Yoshida & Gotoh, 1975). However, the translocation paths of photosynthates have not yet been investigated. Therefore, this study aimed to elucidate the translocation paths and destinations of photosynthates in soybean, using ¹⁴C and CF imaging analysis. As a result, we summarized and updated the model for the translocation and destination of photosynthates in soybean plants (Fig. 6b).

In our initial analysis, we examined the distribution of ¹⁴C-labeled carbon assimilates, which are thought to be photosynthates produced from ¹⁴CO₂ gas in L1. This was subsequently compared to the distribution of ¹⁴C derived from ¹⁴C-sucrose applied to L1. The results showed a similar distribution pattern (Fig. 1). Sucrose is recognized as the primary compound involved in photosynthate translocation in many plants, including soybean(Fisher, 1970; Vernon & Aronoff, 1952). Our experiment thus confirmed that applying ¹⁴C-sucrose to the leaf surface effectively simulates the movement of endogenous photosynthates from the same leaf.

To distinguish the translocation paths of photosynthates from different sources within the same plant, imaging was performed using ¹⁴C-sucrose in combination with CFDA. Initially, ¹⁴C-sucrose and CFDA were added simultaneously to determine whether the ¹⁴C signal from ¹⁴C-sucrose and the fluorescence signal of CF from CFDA exhibited equivalent behavior in the stem (Fig. 3). The results indicated that ¹⁴C and CF were similarly distributed around the cambium in the stem and petiole. Grignon et al. reported that both ¹⁴C-sucrose and CF, when added to the same source leaves of soybean, were translocated to the same sinks, implying that they may pass through common pathways(Grignon et al., 1989). In this study, we were able to directly confirm that ¹⁴C and CF run through the same regions of vascular bundles by imaging techniques. In the cross-section of the stem near the L1 petiole connection, the ¹⁴C and CF signal intensities showed three peaks (Figs. 3c-iii, 5b-ii, iii, S1b-iii and S2b). This is likely because in soybean plants, the leaf vasculature is connected to three wide bundles that enter the stem(Thaine et al., 1959). Above L3 near the stem apex, CF signals were detected solely in the phloem, while ¹⁴C signals were detected in the phloem, xylem, and epidermis (Fig. 3c-e). Since CF moves between cells via plasmodesmata(Jiang et al., 2019), ¹⁴C was thought to be unloaded via pathways other than plasmodesmata. Biddulph et al. and Jones et al. also reported that ¹⁴C was detected in the xylem of young stems when ¹⁴CO₂was absorbed by source leaves(S. Biddulph et al., 1958; Jones et al., 1959). Subsequent imaging experiments were carried out only with mature stems, and ¹⁴C was detected outside the vascular bundles in several cases (Figs. 5b, S1b and S2b, red arrowheads). Although the phloem in the stem is a pathway that translocates photosynthates, the stem also requires nutrients for its own growth and maintenance. It is therefore possible that ¹⁴C was translocated to the outside of the phloem and used as a cell wall component or material for energy production.

This imaging method using a radioisotope and fluorescence is a useful way to compare translocated signals within the same sample. Biddulph et al. previously investigated the translocation paths from different leaf positions in red kidney bean seedlings(O. Biddulph & Cory, 1960). They added ¹⁴CO₂ and NaH₂³²PO₄ to L1 and L2, respectively, and detected radioisotope activity at the L1 to L2 internode. However, these authors were unable to distinguish clearly between the ¹⁴C and ³²P signals(O. Biddulph & Cory, 1960). Furthermore, they also added ¹⁴CO₂ to L1 or L2 of individual plants and compared each ¹⁴C signal in each vascular bundle. Their conclusion was that vascular bundles involved in L1-derived translocation differed from those involved in L2-derived translocation (O. Biddulph & Cory, 1965). In our study, we validated their results visually in cross sections of the same samples (Figs. 5 and S1).

Our imaging analysis revealed photosynthate translocation to young sink leaves from source leaves on opposite side of the stem (Fig. 6b). Photosynthates translocated from L1 and L2 passed through different regions of vascular bundles in the stem but were unloaded at the same destinations, L3 and L4 (Figs. 2, 5 and S1). How is the photosynthate translocated to the sink on the opposite orthostichy? The ¹⁴C and CF intensity graphs showed three peaks near the petiole of leaves to which ¹⁴C-sucrose and CFDA were added (Figs. 3c-iii, 5b-ii, iii, S1b-iii and S2b) but not at a distance. In particular, the upper position in the stem showed two peaks (Figs. 3c-ii, 5b-i, and S1b-i). In the upper stem near the apex, lateral transport may occur within the vascular bundle, expanding the area over which photosynthates are translocated. The ¹⁴C and CF signal peaks from different leaves did not overlap but were not completely separated in the stem section (Figs. 5, S1 and S2). It is also possible that organs with high sink activity may preferentially receive photosynthates from leaves at both the same and opposite orthostichies. Further research is expected to clarify the relationship between sink activity and translocation paths.

In this study, we detected uneven ¹⁴C activity in one leaf. The destination and translocation paths differ depending on whether the photosynthate is derived from the left or right side of the source leaf (Fig. 2c). The translocation paths within the petiole and stem were divided depending on whether ¹⁴C-sucrose was added to the left or right side of the trifoliate (Fig. 4). These results show that photosynthates loaded on the left and right sides of the trifoliate were translocated without mixing and were uniform within a single leaf (Fig. 6b). Uneven ¹⁴C activity in one leaf was also reported in Cucurbita pepo, sugar beet and tobacco, and ¹⁴C-assimilates from source leaves were detected at the leaf base but not at the leaf tip (Fellows & Geiger, 1974; Turgeon & Webb, 1973). This pattern occurs along the distal- proximal axis and is due to different growth stages within a single leaf, so the mechanism is different from the left-right asymmetry observed here. In many other previous experiments with ¹⁴C, although a few autoradiograph images showed uneven distribution in some leaves, the pattern was not regular, and most authors did not mention the distribution. To our knowledge, this is the first report that elucidates the pattern of left-right asymmetric translocation from leaves and its translocation paths.

The findings of this study deepen our understanding of the destination and translocation paths of photosynthates in soybean plants, which can be used to grow soybean plants more efficiently. Furthermore, detailed studies focusing on the reproductive growth stage are needed to enhance yields.

Acknowledgment

This work was supported by JSPS KAKENHI Grant Number JP20H00437. We are grateful to Masako Katsuno for assistance.

Competing Interests

The authors have no conflicts of interest to declare that are relevant to the content of this article.

Alan B. Bennett, Barry L. Sweger, & Roger M. Spanswick. (1984). Sink to Source Translocation in Soybean. Plant Physiology, 74, 434–436.
Biddulph, O., & Cory, R. (1960). Demonstration of Two Translocation Mechanisms in Studies of Bidirectional Movement. Plant Physiology, 35(5), 689–695. https://doi.org/10.1104/pp.35.5.689
Biddulph, O., & Cory, R. (1965). Translocation of C14 Metabolites in the Phloem of the Bean Plant. Plant Physiology, 40(1), 119–129. https://doi.org/10.1104/pp.40.1.119
Biddulph, S., Biddulph, O., & Cory, R. (1958). Visual Indications of Upward Movement of Foliar-Applied P32 and C14 in the Phloem of the Bean Stem. American Journal of Botany, 45(8), 648–652. https://doi.org/10.1002/j.1537-2197.1958.tb10597.x
Burroughs, C. H., Montes, C. M., Moller, C. A., Mitchell, N. G., Michael, A. M., Peng, B., Kimm, H., Pederson, T. L., Lipka, A. E., Bernacchi, C. J., Guan, K., & Ainsworth, E. A. (2023). Reductions in leaf area index, pod production, seed size, and harvest index drive yield loss to high temperatures in soybean. Journal of Experimental Botany, 74(5), 1629–1641. https://doi.org/10.1093/jxb/erac503
Drag, D. W., Slattery, R., Siebers, M., DeLucia, E. H., Ort, D. R., & Bernacchi, C. J. (2020). Soybean photosynthetic and biomass responses to carbon dioxide concentrations ranging from pre-industrial to the distant future. Journal of Experimental Botany, 71(12), 3690–3700. https://doi.org/10.1093/jxb/eraa133
Fellows, R. J., & Geiger, D. R. (1974). Structural and Physiological Changes in Sugar Beet Leaves during Sink to Source Conversion. Plant Physiology, 54(6), 877–885. https://doi.org/10.1104/pp.54.6.877
Feng, L., Raza, M. A., Li, Z., Chen, Y., Khalid, M. H. Bin, Du, J., Liu, W., Wu, X., Song, C., Yu, L., Zhang, Z., Yuan, S., Yang, W., & Yang, F. (2019). The Influence of Light Intensity and Leaf Movement on Photosynthesis Characteristics and Carbon Balance of Soybean. Frontiers in Plant Science, 9. https://doi.org/10.3389/fpls.2018.01952
Fisher, D. B. (1970). Kinetics of C-14 Translocation in Soybean. Plant Physiology, 45(2), 107–113. https://doi.org/10.1104/pp.45.2.107
Grignon, N., Touraine, B., & Durand, M. (1989). 6(5)Carboxyfluorescein as a Tracer of Phloem Sap Translocation. American Journal of Botany, 76(6), 871. https://doi.org/10.2307/2444542
Hartman, G. L., West, E. D., & Herman, T. K. (2011). Crops that feed the World 2. Soybean—worldwide production, use, and constraints caused by pathogens and pests. Food Security, 3(1), 5–17. https://doi.org/10.1007/s12571-010-0108-x
Hitz, T., Hartung, J., Graeff-Hönninger, S., & Munz, S. (2019). Morphological Response of Soybean (Glycine max (L.) Merr.) Cultivars to Light Intensity and Red to Far-Red Ratio. Agronomy, 9(8), 428. https://doi.org/10.3390/agronomy9080428
Jiang, M., Deng, Z., White, R. G., Jin, T., & Liang, D. (2019). Shootward Movement of CFDA Tracer Loaded in the Bottom Sink Tissues of Arabidopsis. Journal of Visualized Experiments, 2019(147). https://doi.org/10.3791/59606
Jones, H., Martin, R. V., & Porter, H. K. (1959). Translocation of 14Carbon in Tobacco following Assimilation of 14Carbon Dioxide by a Single Leaf. Annals of Botany, 23(4), 493–508. https://doi.org/10.1093/oxfordjournals.aob.a083673
Kagawa, T., & Wong, J. H. H. (1985). Allocation and Turnover of Photosynthetically Assimilated 14CO 2in Leaves of Glycine max L. Clark. Plant Physiology, 77(2), 266–274. https://doi.org/10.1104/pp.77.2.266
Kappes, E. M., & Flore, J. A. (1989). Phyllotaxy and Stage of Leaf and Fruit Development Influence Initiation and Direction of Carbohydrate Export from Sour Cherry Leaves. Journal of the American Society for Horticultural Science, 114(4), 642–648. https://doi.org/10.21273/JASHS.114.4.642
Kölling, K., Müller, A., Flütsch, P., & Zeeman, S. C. (2013). A device for single leaf labelling with CO2 isotopes to study carbon allocation and partitioning in Arabidopsis thaliana. Plant Methods, 9(1), 45. https://doi.org/10.1186/1746-4811-9-45
Landau, C. A., Hager, A. G., & Williams, M. M. (2022). Deteriorating weed control and variable weather portends greater soybean yield losses in the future. Science of The Total Environment, 830, 154764. https://doi.org/10.1016/j.scitotenv.2022.154764
Liu, S., Zhang, M., Feng, F., & Tian, Z. (2020). Toward a “Green Revolution” for Soybean. Molecular Plant, 13(5), 688–697. https://doi.org/10.1016/j.molp.2020.03.002
Shishido, Y., & Hori, Y. (1977). Studies on Translocation and Distribution of Photosynthetic Assimilates in Tomato Plants II. Distribution Pattern as Affected by Phyllotaxis. Tohoku Journal of Agricultural Research, 28(2), 82–95.
Shishido, Y., & Hori, Y. (1991). The Role of Leaf as Affected by Phyllotaxis and Leaf Histology on the Development of the Fruit in Tomato. Engei Gakkai Zasshi, 60(2), 319–327. https://doi.org/10.2503/jjshs.60.319
Shishido, Y., Seyama, N., & Hori, Y. (1988). Studies on Distribution Pattern of 14C-assimilates in relation to Vascular Pattern derived from Phyllotaxis of Tomato Plants. Journal of the Japanese Society for Horticultural Science, 57(3), 418–425. https://doi.org/10.2503/jjshs.57.418
Singh, S. K., Reddy, V. R., Devi, M. J., & Timlin, D. J. (2021). Impact of water stress under ambient and elevated carbon dioxide across three temperature regimes on soybean canopy gas exchange and productivity. Scientific Reports, 11(1), 16511. https://doi.org/10.1038/s41598-021-96037-9
Thaine, R., Ovenden, S., & Turner, J. (1959). Translocation of Labelled Assimilates in the Soybean. Australian Journal of Biological Sciences, 12(4), 349–372. https://doi.org/10.1071/BI9590349
Thrower, S. L. (1962). Translocation of labelled assimilates in the soybean. 2. The pattern of translocation in intact and defoliated plants. Aust. J. Biol. Sci., 15, 629–649. https://cir.nii.ac.jp/crid/1571417124622355456.bib?lang=ja
Turgeon, R., & Webb, J. A. (1973). Leaf development and phloem transport in Cucurbita pepo: Transition from import to export. Planta, 113(2), 179–191. https://doi.org/10.1007/BF00388202
Vernon, L. P., & Aronoff, S. (1952). Metabolism of soybean leaves. IV. Translocation from soybean leaves. Archives of Biochemistry and Biophysics, 36(2), 383–398. https://doi.org/10.1016/0003-9861(52)90424-4
Yoshida, K., & Gotoh, K. (1975). Translocation and Distribution of 14C-Assimilates Related to Stem Termination Habits in Soybeans. Japanese Journal of Crop Science, 44(2), 185–193. https://doi.org/10.1626/jcs.44.185
Yoshikawa, T. (2019). The molecular-breeding study of leaf molphogenesis and juvenile-adult phase transition in crops. Breeding Research, 21(2), 150–155. https://doi.org/10.1270/jsbbr.19j12

SIKaihoSoma.pdf

Download PDF

Version 1

posted

You are reading this latest preprint version

Determination of the Phloem Path and Destination of the Photosynthates by Autoradiography and Fluorescent Marker Imaging

Status:

Version 1

Abstract

Figures

Introduction

Materials And Methods

Results

Discussion

Declarations

References

Supplementary Files

Status:

Version 1