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PubMed1991

Developmental and environmental regulation of pea legumin genes in transgenic tobacco.

Two distinct legumin genes (LegA1 and LegA2) which encode a major class of seed storage protein in pea were isolated from a genomic library. The cloned fragments were introduced into tobacco via Agrobacterium-mediated transformation and the regenerated plants were used to study the expression characteristics of the genes in a heterologous host. It was found that both LegA1 and LegA2 were functional members of the pea legumin gene family and that their expression was similar in both pea and transgenic tobacco. Legumin was detected only in the seed of tobacco where the primary translation products were processed in a manner analogous to that which occurs in pea. Legumin gene expression was also shown to be temporally regulated during seed development. Legumin polypeptides and mRNA began to accumulate 16 days after flowering (DAF), in contrast to the endogenous tobacco storage proteins which were apparent at 13 DAF. It was also demonstrated that the legumin genes in tobacco were environmentally regulated to the nutritional status of the plant. As has been previously shown in pea, legumin accumulation in transgenic tobacco seed was progressively reduced when the plants were grown under conditions of increasing severity of sulphur-nutrient stress. The reduced accumulation of protein was correlated with lower levels of legumin mRNA in the developing seed. Despite encoding nearly identical subunits, nucleotide sequence data for LegA1 and LegA2 showed that the similarity of their respective 5'-flanking regions was restricted to several short elements mostly within 240 bp from the start of transcription. However, a deletion series using the LegA1 gene demonstrated that 237 bp of 5'-flanking sequence was insufficient to permit the expression of the legumin gene in tobacco. The data indicated that an as yet unidentified sequence element(s) located between positions -668 and -237 was essential in re-establishing the high level of regulated gene expression observed with the full-length LegA1 gene.

Base Sequence↗

PubMed2004

Thioredoxin targets of developing wheat seeds identified by complementary proteomic approaches.

The role of thioredoxin in wheat starchy endosperm was investigated utilizing two proteomic approaches. Thioredoxin targets were isolated from total KCl-soluble extracts of endosperm and flour and separated by 2-DE following (1) reduction of the extract by the NADP/thioredoxin system and labeling the newly generated sulfhydryl (SH) groups with monobromobimane (mBBr), and, in parallel, (2) trapping covalently interacting proteins on an affinity column prepared with mutant thioredoxin h in which one of the active site cysteines was replaced by serine. The two procedures were complementary: of the total targets, one-third were observed with both procedures and one-third were unique to each. Altogether 68 potential targets were identified; almost all containing conserved cysteines. In addition to confirming known interacting proteins, we identified 40 potential thioredoxin targets not previously described in seeds. A comparison of the results obtained with young endosperm (isolated 10 days after flowering) to those with mature endosperm (isolated 36 days after flowering) revealed a unique set of proteins functional in processes characteristic of each developmental stage. Flour contained 36 thioredoxin targets, most of which have been found in the isolated developing endosperm.

Bridged Bicyclo Compounds↗

PubMed2003

Pea seed-borne mosaic virus seed transmission exploits novel symplastic pathways to infect the pea embryo and is, in part, dependent upon chance.

Seed transmission of pea seed-borne mosaic virus (PSbMV) depends upon symplastic transport of the virus from infected maternal cells to the embryo. Such transport pathways have not been identified in higher plants. To identify these pathways, we have studied the ultrastructure of the tissues and cells around the micropyle of young developing seeds and compared transmitted and nontransmitted virus isolates. A characteristic of PSbMV infection was the presence of cylindrical inclusions positioned over plasmodesmal openings. The presence of cylindrical inclusions on the testa-endosperm boundary wall, together with immunogold labelling for virus-specific products on the wall and in the endosperm, indicated that symplastic connections existed at this interface. Close examination of the endosperm-suspensor boundary at the base of the suspensor revealed discontinuities in the suspensor sheath wall as porelike structures, which the virus might pass through en route to the embryo. A nontransmitted PSbMV isolate was able to invade the maternal tissues of the developing seed but was excluded from the embryo, although it was detected at a low level in the endosperm. Since the endosperm did not support virus replication, it appeared that passive accumulation determined the amount, timing, and location of the virus relative to the base of the suspensor. Rarely, therefore, could the nontransmitted virus isolate reach the correct location in the endosperm at the correct time for embryo infection via the suspensor to occur.

Embryo, Mammalian↗

PubMed2002

Distinct physiological roles of fructokinase isozymes revealed by gene-specific suppression of Frk1 and Frk2 expression in tomato.

There are two divergent fructokinase isozymes, Frk1 and Frk2 in tomato (Lycopersicon esculentum Mill.) plants. To investigate the physiological functions of each isozyme, the expression of each fructokinase mRNA was independently suppressed in transgenic tomato plants, and the respective phenotypes were evaluated. Suppression of Frk1 expression resulted in delayed flowering at the first inflorescence. Suppression of Frk2 did not effect flowering time but resulted in growth inhibition of stems and roots, reduction of flower and fruit number, and reduction of seed number per fruit. Localization of Frk1 and Frk2 mRNA accumulation by in situ hybridization in wild-type tomato fruit tissue indicated that Frk2 is expressed specifically in early tomato seed development. Fruit hexose and starch content were not effected by the suppression of either Frk gene alone. The results collectively indicate that flowering time is specifically promoted by Frk1 and that Frk2 plays specific roles in contributing to stem and root growth and to seed development. Because Frk1 and Frk2 gene expression was suppressed individually in transgenic plants, other significant metabolic roles of fructokinases may not have been observed if Frk1 and Frk2 play, at least partially, redundant metabolic roles.

Enzyme Activation↗

PubMed2006

Biochemical and genomic analysis of sucrose metabolism during coffee (Coffea arabica) fruit development.

Sucrose metabolism and the role of sucrose synthase were investigated in the fruit tissues (pericarp, perisperm, and endosperm) of Coffea arabica during development. Acid invertase, sucrose phosphate synthase, and sucrose synthase activities were monitored and compared with the levels of sucrose and reducing sugars. Among these enzymes, sucrose synthase showed the highest activities during the last stage of endosperm and pericarp development and this activity paralleled closely the accumulation of sucrose in these tissues at this stage. Carbon partitioning in fruits was studied by pulse-chase experiments with (14)C-sugars and revealed high rates of sucrose turnover in perisperm and endosperm tissues. Additional feeding experiments with (14)CO(2) showed that leaf photosynthesis contributed more to seed development than the pericarp in terms of photosynthate supply to the endosperm. Sugar analysis, feeding experiments, and histological studies indicated that the perisperm plays an important role in this downloading process. It was observed that the perisperm presents a transient accumulation of starch which is degraded as the seed develops. Two full-length cDNAs (CaSUS1 and CaSUS2) and the complete gene sequence of the latter were also isolated. They encode sucrose synthase isoforms that are phylogenetically distinct, indicating their involvement in different physiological functions during cherry development. Contrasting expression patterns were observed for CaSUS1 and CaSUS2 in perisperm, endosperm, and pericarp tissues: CaSUS1 mRNAs accumulated mainly during the early development of perisperm and endosperm, as well as during pericarp growing phases, whereas those of CaSUS2 paralleled sucrose synthase activity in the last weeks of pericarp and endosperm development. Taken together, these results indicate that sucrose synthase plays an important role in sugar metabolism during sucrose accumulation in the coffee fruit.

Biological Transport↗

PubMed2001

LEAFY COTYLEDON2 encodes a B3 domain transcription factor that induces embryo development.

The Arabidopsis LEAFY COTYLEDON2 (LEC2) gene is a central embryonic regulator that serves critical roles both early and late during embryo development. LEC2 is required for the maintenance of suspensor morphology, specification of cotyledon identity, progression through the maturation phase, and suppression of premature germination. We cloned the LEC2 gene on the basis of its chromosomal position and showed that the predicted polypeptide contains a B3 domain, a DNA-binding motif unique to plants that is characteristic of several transcription factors. We showed that LEC2 RNA accumulates primarily during seed development, consistent with our finding that LEC2 shares greatest similarity with the B3 domain transcription factors that act primarily in developing seeds, VIVIPAROUS1/ABA INSENSITIVE3 and FUSCA3. Ectopic, postembryonic expression of LEC2 in transgenic plants induces the formation of somatic embryos and other organ-like structures and often confers embryonic characteristics to seedlings. Together, these results suggest that LEC2 is a transcriptional regulator that establishes a cellular environment sufficient to initiate embryo development.

Amino Acid Sequence↗

PubMed2001

Developmental variation of sugars, carboxylic acids, purine alkaloids, fatty acids, and endoproteinase activity during maturation of Theobroma cacao L. seeds.

The changes of mono- and oligosaccharides, carboxylic acids, purine alkaloids, and fatty acid composition, and of aspartic endoproteinase activity, were analyzed during seed development in two varieties of cacao (Theobroma cacao). The majority of the components examined either decreased or accumulated steadily in concentration during the second half of bean development. Sucrose is the major sugar in the mature embryo, whereas fructose and glucose are at higher concentrations in the endosperm tissue. Considerable amounts of malate are found in the endosperm, whereas citrate is the dominant carboxylic acid in the embryo. A major change in the fatty acid composition occurs in the young embryo when the proportion of stearic acid increases rapidly at the expense of linoleic acid, which is reduced from about 18 to 3%. Theobromine is the dominant purine alkaloid (ca. 80%), and caffeine appears only toward the end of seed maturity. Aspartic endoproteinase activity increases rapidly during embryo expansion, reaching a maximal activity before final maturity. The results are discussed in conjunction with physiological changes in developing seeds, and the potential contributions of the compounds analyzed for cocoa quality.

Aspartic Acid Endopeptidases↗

PubMed2002

Genomic organization of glycinin genes in soybean.

Glycinin is the predominant seed storage protein in most soybean varieties. Previously, five major genes (designated Gy1 to Gy5) encoding glycinin subunits have been described. In this report two new genes are identified and mapped: a glycinin pseudogene, gy6, and a functional gene, Gy7. Messenger RNA for the gy6 pseudogene is not detected in developing seeds. While Gy7 mRNA was present at the midmaturation stage of seed development in the soybean variety Resnik, the steady state amount of this message was at least an order of magnitude less-prevalent than the mRNA encoding each of the other five glycinin subunits. Even though the amino-acid sequence of the glycinin subunit G7 is related to the other five soybean 11S subunits, it does not fit into either the Group-1 (G1, G2, G3) or the Group-2 (G4, G5) glycinin subunit families. The Gy7 gene is tandemly linked 3' to Gy3 on Linkage Group L (chromosome 19) of the public molecular linkage map. By contrast, the gy6 gene occupies a locus downstream from Gy2 on Linkage Group N (chromosome 3) in a region that is related to the position where Gy7 is located on chromosome 19.

Journal Article↗

PubMed2004

Development of an efficient method for the isolation of factors involved in gene transcription during rice embryo development.

Summary An efficient yeast-based system was developed for the isolation of plant cDNAs encoding transcription factors (TFs) and proteins with transcription activation functions (co-activators). The system consists of two vectors: (i) a reporter vector (pG221) harboring the iso-1-cytochrome c (CYC1) core promoter and the beta-galactosidase (lacZ) gene; and (ii) a cDNA library construction vector (pYF503), which yields a library of plant peptides fused to the GAL4-binding domain (GAL4-BD). Expression of a peptide harboring the characteristics of a transcriptional activator leads to expression of lacZ, allowing for selection of relevant colonies. TFs during rice embryo development were isolated through this system. Approximately 200 confirmed positive colonies were obtained from screening 10(6) yeast colonies, and sequence analysis of conserved domains identified 75 independent cDNAs, 20 of which encoded plant TFs or co-activators, including members of the APETALA2 (AP2)/ethylene-responsive element-binding protein (EREBP), MYB and growth-regulating factor (GRF) families. Peptides encoded by 13 of the isolated cDNAs were classified as potential TFs or co-activators because of the presence of conserved TF-like domains. Additionally, 2, 11, and 13 clones encoded kinases, chromosome-related proteins, and unknown proteins, respectively, while the remaining 16 cDNAs were associated with specific functions seemingly unrelated to TFs. Expression pattern analysis of selected TF-encoding genes via RT-PCR revealed that these genes were expressed during seed development, with differential transcription observed during various stages. This work provides informative hints for further study of the regulatory mechanism of rice seed development and illustrates an identification strategy that will be of practical value for the isolation of TFs and co-activators associated with specific plant developmental processes.

Amino Acid Sequence↗

PubMed2000

Characterisation of acyl-ACP desaturases from Macadamia integrifolia Maiden & Betche and Nerium oleander L.

The seed oil in Macadamia integrifolia contains about 30% palmitoleic acid (16:1(Delta9)) and Nerium oleander about 12% isoricinoleic acid (Delta9-hydroxy-18:1(Delta12)). It has been shown that palmitoleic acid can be produced by acyl-acyl carrier protein (ACP) desaturases and it has also been shown that fatty acid hydroxylation can occur via direct substitution of a hydrogen atom. Therefore it seemed possible that the enzymes responsible for the making of these unusual fatty acids in M. integrifolia and N. oleander were of acyl-ACP desaturase type. Extracts from developing M. integrifolia developing seeds showed a relative ratio of 16:0-ACP to 18:0-ACP desaturation that was about 13 times higher than in sunflower seeds. N. oleander seed extracts catalysed conversion of 18:0-ACP to 18:1(Delta9) but only trace amounts of Delta9-hydroxy fatty acids were formed. A total of four cDNAs were isolated from developing seeds, of both species, using a fragment isolated with PCR amplification. The M. integrifolia acyl-ACP desaturase cDNA was expressed in Escherichia coli. A partly purified fraction of the enzyme showed a 16:0-ACP to 18:0-ACP desaturation ratio about 90-fold less than that in the Macadamia extracts. Expressed N. oleander acyl-ACP desaturase cDNAs showed predominantly 18:0-ACP desaturase activity and no hydroxylase activity. Thus it is not likely that any of the four acyl-ACP desaturases cloned from M. integrifolia or N. oleander is involved in the production of unusual fatty acids.

Journal Article↗

PubMed1985

Effects of nutritional stress on the storage proteins of soybeans.

The effects of sulfur deficiency on the complement of proteins laid down in developing seeds of soybean (Glycine max L. Merr) have been examined. Sulfur deficiency caused a 40% decrease in the level of glycinins and a contrasting elevation in the level of beta-conglycinins. The subunit composition of these proteins was also affected. There was in particular a 3-fold increase in the beta-subunit of beta-conglycinins in the sulfur-deficient seeds, and this accumulated largely as the B(0)-isomer of beta-conglycinins, a protein which while virtually devoid of methionine and cysteine retains the physical properties of a normal 7S storage protein. These data demonstrate that a high degree of selectivity can be exerted by environmental stress over the accumulation of proteins in developing seeds.

Journal Article↗

PubMed1992

Molecular cloning of a gene encoding a seed-specific protein with nutritionally balanced amino acid composition from Amaranthus.

An albumin with a well-balanced amino acid composition and high levels of the essential amino acids was purified to homogeneity from the mature seeds of Amaranthus hypochondriacus. The amino acid composition of this protein is comparable to the World Health Organization recommended values for a highly nutritional protein. The protein is a 35-kDa monomer with four isoforms that can be separated by chromatofocusing. Antibodies raised against one of the isoforms, AmA1, cross-reacted with the other three isoforms. Affinity-purified AmA1 antibodies were used to isolate cDNA clones from a developing-seed expression library. The six immunopositive recombinants obtained were found to be related. The cDNA of the largest clone (1.2 kilobases) has a single major open reading frame corresponding to a 304-amino acid polypeptide. The clone was confirmed by hybrid-selected translation and immunoprecipitation. The size of the immunoprecipitated product was identical to the mature protein. Analysis of RNA and protein in developing seeds showed that AmA1 is synthesized during early embryogenesis, reaching a maximum by midmaturation. No RNA was detected in 1-day-old seedlings although the protein showed delayed breakdown on germination. Expression of the AmA1 gene was found to be seed-specific, as no protein or RNA was detected in other plant tissues.

Amino Acid Sequence↗

PubMed1998

The biosynthesis of erucic acid in developing embryos of brassica rapa

The prevailing hypothesis on the biosynthesis of erucic acid in developing seeds is that oleic acid, produced in the plastid, is activated to oleoyl-coenzyme A (CoA) for malonyl-CoA-dependent elongation to erucic acid in the cytosol. Several in vivo-labeling experiments designed to probe and extend this hypothesis are reported here. To examine whether newly synthesized oleic acid is directly elongated to erucic acid in developing seeds of Brassica rapa L., embryos were labeled with [14C]acetate, and the ratio of radioactivity of carbon atoms C-5 to C-22 (de novo fatty acid synthesis portion) to carbon atoms C-1 to C-4 (elongated portion) of erucic acid was monitored with time. If newly synthesized 18:1 (oleate) immediately becomes a substrate for elongation to erucic acid, this ratio would be expected to remain constant with incubation time. However, if erucic acid is produced from a pool of preexisting oleic acid, the ratio of 14C in the 4 elongation carbons to 14C in the methyl-terminal 18 carbons would be expected to decrease with time. This labeling ratio decreased with time and, therefore, suggests the existence of an intermediate pool of 18:1, which contributes at least part of the oleoyl precursor for the production of erucic acid. The addition of 2-[3-chloro-5-(trifluromethyl)-2-pyridinyloxyphenoxy] propanoic acid, which inhibits the homodimeric acetyl-CoA carboxylase, severely inhibited the synthesis of [14C]erucic acid, indicating that essentially all malonyl-CoA for elongation of 18:1 to erucate was produced by homodimeric acetyl-CoA carboxylase. Both light and 2-[3-chloro-5-(trifluromethyl)-2-pyridinyloxyphenoxy]-propanoic acid increased the accumulation of [14C]18:1 and the parallel accumulation of [14C]phosphatidylcholine. Taken together, these results show an additional level of complexity in the biosynthesis of erucic acid.

Journal Article↗

PubMed1999

Molecular characterization of a gene for alanine aminotransferase from rice (Oryza sativa).

A cDNA clone encoding alanine aminotransferase (AlaAT) has isolated from randomly sequenced clones derived from a cDNA library of maturing rice seeds by comparison to previously identified genes. The deduced amino acid sequence was 88% and 91% homologous to those of the enzymes from barley and broomcorn millet (Panicum miliaceum), respectively. Using this cDNA as a probe, we isolated and sequenced the corresponding genomic clone. Comparison of the sequences of the cDNA and the genomic gene revealed that the coding region of the gene was interrupted by 14 introns 66 to 1547 bp long. Northern and western blotting analyses showed that the gene was expressed at high levels in developing seeds. When the 5'-flanking region between -930 and +85 from the site of initiation of transcription was fused to a reporter gene for beta-glucuronidase (GUS) and then introduced into the rice genome, histochemical staining revealed strong GUS activity in the inner endosperm tissue of developing seeds and weak activity in root tips. Similar tissue-specific expression was also detected by in situ hybridization. These results suggest that AlaAT is involved in nitrogen metabolism during the maturation of rice seed.

Alanine Transaminase↗

PubMed1989

Regulated expression of genes encoding soybean beta-conglycinins in transgenic plants.

Beta-conglycinin, an abundant storage protein in soybean seeds, is comprised of three subunits: alpha', alpha, and beta. Expression of genes encoding these subunits is tissue-specific and temporally regulated in soybean plants. Both the alpha'- and beta-subunit genes have been isolated and fully or partially characterized by DNA sequence analysis. Each has been transferred to transgenic plants and was found to be expressed in petunia and tobacco plants in a regulated manner as in soybean plants. Both the alpha'- and beta-subunit proteins were accumulated only in transgenic seeds during mid-to-late stages of seed development and assembled into multimeric forms with sedimentation coefficients of 7S and 9S. The alpha'- and beta-subunit genes were also introduced on the same plasmid into transgenic plants. When the promoters of the two genes were proximal to each other, there was an increase in the expression of the beta-subunit gene, suggesting that there is a strong cis-acting sequence in the alpha'-gene (promoter) that can affect expression of the beta-subunit gene. A series of deletion mutants was constructed in the alpha'-gene promoter, and each was introduced into petunia plants. The DNA sequence approximately 250 bp upstream of the transcriptional initiation site contains sufficient regulatory information for expression of the alpha'-subunit gene. The sequence about 200 bp upstream contains four 6-bp repeats A (AGCCCA) that may play an important role in regulating expression of the alpha'-gene. The function of this DNA sequence (-78 to -257) was tested in a chimeric gene construct containing the CaMV 35S promoter, the CAT coding sequence, and the 3' noncoding region of the alpha'- gene. A dramatic enhancement of the CAT gene expression (at least 40-fold) resulted when this DNA sequence was inserted into the 35S promoter at a position 90 bp upstream of the transcription start site, regardless of the orientation of the insert. There was a two- to threefold enhancement when the element was placed 6' to the CAT coding region and no enhancement when placed downstream of the 3' noncoding region. The enhancement was observed only in seeds during mid-to-late stages of seed development. This indicates that the 170-bp DNA sequence can function as a strong cis-acting element that controls seed-specific and temporally regulated gene expression. This element is defined as a seed-specific and developmentally regulated element (SDRE).

Antigens, Plant↗

PubMed2005

Genetic mapping and comparative analysis of seven mutants related to seed fiber development in cotton.

Mapping of genes that play major roles in cotton fiber development is an important step toward their cloning and manipulation, and provides a test of their relationships (if any) to agriculturally-important QTLs. Seven previously identified fiber mutants, four dominant (Li (1), Li (2), N (1) and Fbl) and three recessive (n (2), sma-4(h (a)), and sma-4(fz)), were genetically mapped in six F(2) populations comprising 124 or more plants each. For those mutants previously assigned to chromosomes by using aneuploids or by linkage to other morphological markers, all map locations were concordant except n (2), which mapped to the homoeolog of the chromosome previously reported. Three mutations with primary effects on fuzz fibers (N (1), Fbl, n (2)) mapped near the likelihood peaks for QTLs that affected lint fiber productivity in the same populations, perhaps suggesting pleiotropic effects on both fiber types. However, only Li (1) mapped within the likelihood interval for 191 previously detected lint fiber QTLs discovered in non-mutant crosses, suggesting that these mutations may occur in genes that played early roles in cotton fiber evolution, and for which new allelic variants are quickly eliminated from improved germplasm. A close positional association between sma-4(h ( a )), two leaf and stem-borne trichome mutants (t (1) , t (2)), and a gene previously implicated in fiber development, sucrose synthase, raises questions about the possibility that these genes may be functionally related. Increasing knowledge of the correspondence of the cotton and Arabidopsis genomes provides several avenues by which genetic dissection of cotton fiber development may be accelerated.

Chromosome Mapping↗

PubMed2000

ABI3 affects plastid differentiation in dark-grown Arabidopsis seedlings.

The Arabidopsis ABSCISIC ACID-INSENSITIVE3 (ABI3) protein has been identified previously as a crucial regulator of late seed development. Here, we show that dark-grown abi3 plants, or abi3 plants returned to the dark after germination in the light, developed and maintained an etioplast with a prominent prolamellar body at developmental stages in which the wild type did not. Overexpression of ABI3 led to the preservation of the plastid ultrastructure that was present at the onset of darkness. These observations suggest that ABI3 plays a role in plastid differentiation pathways in vegetative tissues. Furthermore, the analysis of deetiolated (det1) abi3 double mutants revealed that DET1 and ABI3 impinge on a multitude of common processes. During seed maturation, ABI3 required DET1 to achieve its full expression. Mature det1 abi3 seeds were found to be in a highly germinative state, indicating that germination is controlled by both DET1 and ABI3. During plastid differentiation in leaves of dark-grown plants, DET1 is required for the action of ABI3 as it is during seed development. Together, the results suggest that ABI3 is at least partly regulated by light.

Arabidopsis↗

PubMed1987

Effect of head removal on leaf senescence of sunflower.

Greenhouse and field studies examined the effect of flower or seedhead removal on leaf senescence and associated changes in sunflower (Helianthus annuus L.) plants. At intervals during seed development, selected leaves (leaves 6 through 8 from the top in the greenhouse and leaf 7 from the top in the field) were harvested and analyzed for chlorophyll, specific leaf weight, N, P, soluble protein, and electrophoretic gel profiles of soluble polypeptides. In both the greenhouse and the field, the leaves of headless plants retained or accumulated more N, P, soluble protein, and dry weight than leaves of plants with heads. Obviously, head removal affected the partitioning of these metabolites during seed development. None of the treatments resulted in the formation of new polypeptides (electrophoretic gel profiles). Comparisons of the rates and extent of loss of chlorophyll, soluble protein, and polypeptide bands (especially ribulose 1,5-bisphosphate carboxylase) from the leaves of headed and deheaded plants showed that head removal delayed the rate of development of leaf senescence for the greenhouse-grown but had much less effect on field-grown plants. These findings illustrate the variability in different parameters commonly associated with the leaf senescence processes of headed and deheaded sunflower plants grown under different environments.

Journal Article↗