Our analysis of Solanaceae euFUL homologs show that FUL1 and FUL2 are broadly expressed in leaves, flowers, and fruit . This overall similarity in expression may indicate a conservation of cis-regulatory elements in gene copies following duplication . Supporting this, our investigation into the number of putative TF binding sites in the promoter region of euFULI homologs did not reveal statistically significant differences . In tomato fruit development, FUL1 expression increases with time, whereas FUL2 expression reaches a maximum at early stages and then decreases over later stages . This variation in expression associated with the developmental stages might be due to changes in cis-elements as a result of the accumulation of random mutations over time . Our analysis did find differences in the number and location of predicted binding sites for specific TFs or families, for instance for ARF, STK, and EIN3 TFs, which may account for the types of differences in expression seen between euFUL paralogs. The 5 kb region upstream of the FUL1 transcription start site in tomato contains three putative ARF binding sites but the corresponding region of FUL2 in tomato contains no such motifs . ARF TFs, raspberry cultivation pot important in tomato fruit development, are activated in response to auxin and may upregulate or repress downstream genes ; the absence of binding sites from the FUL2 promoter is the type of factor that might underlie differences in expression observed between FUL1 and FUL2. Predicted STK binding sites are only found in the promoters of potato FUL1, tomato FUL2 and woodland tobacco MBP10.
STK and STK like proteins appear to function in storage protein synthesis, glucose reception, and vegetative and reproductive development . Meanwhile, the 2 kb upstream region of FUL2 contains a putative site for EIN3. This protein is involved in the development of tomato in response to ripening-associated ethylene production . No such motifs are found in the corresponding region of FUL1. In contrast, the 2–5 kb region in FUL2 contains four putative sites for EIN3 while the corresponding region in FUL1 contains three such sites . Such variation in number and location of TF binding sites has been shown to be associated with the temporal differences in gene expression . Whereas the euFULI members largely overlap in spatial expression with some variation associated with developmental stages, the euFULII homologs show less consistent spatial expression patterns. Only MBP20 is expressed in tomato roots and potato fruit while only MBP10 is expressed in potato tubers . However, these “on” or “off ” expression patterns cannot be explained by the presence or absence of any putative TF binding sites . These two paralogs, which appear to be the result of a tandem duplication and inversion, are located approximately 14.3 Mbp apart on chromosome 2. Although gene clusters resulting from tandem duplications are often coexpressed, this is not the case when there are large physical distances between the genes . An investigation into the expression of human transgenes in mice also found changes in expression as a consequence of an inversion, possibly through disrupting enhancer activity or changes to chromatin structure . Chromosomal rearrangements such as inversions may also result in novel connections between coding regions and other promoters or long distance regulatory motifs while disrupting the original regulatory mechanisms . This sort of re-coupling of one of the two paralogs might lead to the types of contrasting expression patterns observed for MBP10 and MBP20.
However, the expression patterns are not consistent across species and this might be due to additional changes following the inversion . An in-depth analysis of the entire loci and their genomic environment for all paralogs in multiple species would be necessary to determine if the tandem duplication and inversion are associated with changes in proximity to heterochromatin, additional rearrangements, or other phenomena that might have altered gene expression.The first intron of some MADS-box genes contains cis-elements important for the regulation of expression . Studies have found that deletions in the first intron of a FUL-like gene in Aegilops tauschii alters expression and results in the loss of the vernalization requirement . Consistent with this, the first introns of angiosperm euFUL orthologs are generally in the range of 1–10 kb . In contrast, tomato MBP10 has a short first intron of 80 bp. We compared the putative TF binding sites in the first introns of MBP10 and MBP20 in tomato to characterize potential loss of such sites, which might suggest reduced gene regulation. The first intron of MBP10 is predicted to have no TF binding sites, while the first intron of MBP20 is predicted to contain 88 TF binding sites . These included binding sites for MYB, HSF, Dof, WRKY, and MADS-box TFs. Specific TFs predicted to bind to these sites include MYB2 and C1 , which play roles in anthocyanin accumulation and lignin biosynthesis, PBF , which plays a role in endosperm storage protein accumulation, and SPF1 , thought to function in fruit ripening . A similar pattern was found in analysis of the first intron of MBP10 in Nicotiana obtusifolia, which is 110 bp . This analysis found three putative TF binding sites for MYB2 and one for PBF. By contrast, the first intron of N. obtusifolia MBP20 is predicted to have 133 TF binding sites and include a repertoire similar to those found for tomato MBP20.
To determine whether the difference in TF binding site number between the paralogs represented a gain of sites in the MBP20 genes or a loss in the MBP10 genes, we also searched for TF binding sites in the first intron of AGL79, the single euFULII ortholog in A. thaliana . We found that it contains 49 predicted TF binding sites for five different TFs in four families: MYB , HSF , WRKY , and GT-box . Although this number is substantially smaller than the number of sites predicted in the first introns of the Solanaceae MBP20 genes, the results suggest that there has been a loss of TF binding sites in MBP10. Core-eudicot euFUL and basal-eudicot FUL-like genes frequently have broad expression patterns and are generally expressed in fruit . Therefore, the absence or extremely weak expression of MBP10 in fruits of all species, and its weak expression in most organs of tomato and potato is notable . This relatively weak expression may at least in part be due to the loss of TF binding sites in the first intron and suggests a potentially reduced role in regulating fruit-related developmental processes. Importantly, the loss of putative TF binding sites and low expression, combined with the faster evolutionary rate, suggest MBP10 might be in the process of becoming a pseudogene. Further support for this hypothesis comes from an examination of the MBP10 sequences, which suggests that at least two of the sequences in our study show a frame shift that would result in an premature stop codon.There are many ways to train and prune deciduous fruit trees, and no single method is right for all situations and needs. When selecting fruit trees, one important consideration is the desired size of the trees at maturity. Many people prefer small trees because they are easier to manage and harvest and because more trees can be grown in a limited space. Other people prefer full-sized trees because they provide more shade and more fruit per unit area.Genetic dwarf trees usually produce very short internodes , resulting in compact branches with dense foliage. These trees grow to about 8 to 10 feet tall and wide at maturity. They make beautiful landscape trees that are easily managed to provide adequate amounts of fruit for a single family. Excellent varieties are available in peaches, nectarines, and apples. The lower and interior fruiting branches of genetic dwarf trees, especially peaches and nectarines, low round pots tend to die quickly due to shading by the dense growth. However, the trees are small, so production of fruit on the extremities of the higher branches is not a serious problem as long as the branches are strong enough to hold the weight of the fruit. Pruning genetic dwarf trees mainly involves thinning the branches in the dormant season to open up the canopy and maintain the height and spread of the tree. Control tree size and strengthen limbs by removing branches at their point of attachment to the trunk or a larger branch , rather than by heading or “topping” them. For definitions of pruning terms, refer to the glossary.Full-sized trees on standard rootstocks can grow to 25 to 30 feet tall, while trees on semidwarfing rootstock can reach 15 to 20 feet tall. Both standard and semidwarf trees can be kept relatively small by pruning, but trees of this size may still grow too large for many backyard situations. An excellent selection of truly dwarfing apple rootstocks is usually available, and truly dwarfing rootstocks are being developed for most fruit species. Depending on the type of tree , full-sized and semidwarf trees may be trained to an open center, central leader, or fruit bush system.The open center, or vase-shaped, system is most commonly used on almond, apricot, cherry, fig, nectarine, peach, plum, and prune trees. Many pear, apple, and pistachio trees are also trained to this system. With this method, the center of the tree is kept free of large branches and vigorous upright shoots in order to allow sunlight to reach the lower fruiting wood. First growing season.
To create an open center tree, in about late April of the first growing season select three or four shoots that will become the primary scaffold branches and pinch back all other strong upright shoots to 4 to 6 inches long. When possible, these scaffold branches should be spaced several inches apart vertically and should be distributed evenly around the trunk, with the lowest branch about 18 to 24 inches above the ground. If growth is vigorous, the selected scaffold branches should be pinched back or headed to about 2 to 21⁄2 feet long in late May or early June to promote side branching and the development of secondary scaffold branches. Continue to pinch or head back unwanted branches but leave lateral shoots for next year’s fruit production. These unwanted branches will be removed later, but they provide shade for the trunk and main branches of young trees during the current growing season. If summer pruning was not done or was insufficient, create the open center during the dormant season. First dormant season after planting. Select three or four primary scaffold branches if this was not done the previous summer. Do not select scaffold limbs that grow directly above one another. Avoid upright limbs that are attached with narrow, acute angles because they tend to be weak at the point of attachment. Flat or horizontal limbs should be avoided as scaffold limbs, but they can be used if new shoots coming from them are directed upward and outward. For most species, a 45º angle for limb attachment is most desirable. If the tree grows poorly the first year, severely head back the primary scaffolds to three or four buds to promote vigorous growth the next year and correct the causes of the poor growth. Cherry, plum, and pear trees produce very upright growth. To promote tree spread, the scaffolds should be bent outward while still flexible or cut back to outside lateral branches. Other trees, such as apricots, peaches, and almonds, have a spreading growth habit and tend to produce lateral branches without heading. With these varieties it is often necessary to remove flatter-angled branches and leave more upright branches, thus maintaining the upward, outward growth pattern. Developing the mature tree. Heading or pinching the primary scaffold branches encourages secondary scaffold branches to grow from them. Allow two to three secondary branches to develop from each primary. Remove all other strong branches, preferably during the summer, to reduce competition with the scaffold branches and providesunlight to lateral fruiting branches and spurs. Head or pinch the secondary scaffold branches at 2 to 21⁄2 feet long to develop two to three “tertiary” branches from each secondary branch. Ideally, two branches should originate from each primary and secondary, and they should grow upward and outward but away from each other. However, shoot growth seldom conforms to this structure. Mature trees.