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Fan Cao, Xinwang Wang, Zhuangzhuang Liu, Yongrong Li, and Fangren Peng

Pecan cuttings are difficult for rooting. This study describes the pecan hardwood rooting process based on anatomic characteristics to understand root formation mechanisms of pecan cuttings. The expressed proteins of different periods during the adventitious rooting process of pecan seedling hardwood cuttings were identified and analyzed to evaluate the rooting mechanism. The expressed proteins of pecan cutting seedlings were also compared with other cultivar cuttings during the rooting period. Pecan seedling cuttings were developed at different air and substrate temperatures to induce root formation. Adventitious root formation of pecan hardwood cuttings was described, and the phloem at the base of the prepared cuttings was selected as the sample for the differential protein analysis. The results showed that adventitious root formation of pecan hardwood cuttings was the only product of callus differentiation, which originated from the cells of the cambium or vascular ray parenchyma. Such adventitious root primordia were developed from those calluses that formed the regenerative structure, and the expressed proteins during the adventitious rooting of pecan hardwood cutting were identified and analyzed by matrix-assisted laser desorption ionization–time of flight–mass spectrometry (MALDI-TOF-MS) to evaluate the rooting mechanism. Eight differentially expressed proteins were found in the rooting periods, and 15 differential proteins were found by comparing pecan cutting types, which were analyzed by peptide mass fingerprinting homology. The results show that the primordial cells were differentiated from the meristematic cells. Furthermore, the differentially expressed proteins contained energy metabolism proteins, adversity stress proteins, and signal transmission proteins. The energy metabolism-related proteins were adenosine triphosphate (ATP) synthase, photosynthesis-related proteins, and enolase. The adversity-stress proteins containing heat shock-related proteins and signal transmission proteins were mainly cytochrome enzymes and heme-binding proteins. Adventitious root formation of pecan cultivar hardwood cuttings was difficult. More trials should be performed from the potential aspects of high defensive protection and phloem morphologic structure.

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Fan Cao, Yunchu Wei, Xinwang Wang, Yongrong Li, and Fangren Peng

To describe the influence of different pecan seedling rootstocks on drought stress resistance, 12 rootstocks of ‘87MX1-2.2’, ‘87MX5-1.7’, ‘Elliott’, ‘Frutoso’, ‘Giles’, ‘Major’, ‘Moore’, ‘Peruque’, ‘Posey’, ‘Riverside’, ‘San Felipe’, and ‘VC1-68’ were selected as rootstock treatments for grafting. In addition, the experimental materials for the grafted young ‘Pawnee’ tree treatments included the pressure-volume technique (PV technique) and cutting shoot transpiration methods to plot the PV and the cutting shoot transpiration curves, and the parameters calculated from the two curves were used to analyze the data produced by the subordinate function and cluster dendrogram methods. The results revealed that the different seedling rootstock treatments influenced the ‘Pawnee’ grafted trees to varying degrees on aspects of drought resistance, the ability to save water, the modulation of osmosis, and the sensitivity of the stomatal response. The order of drought tolerance for these different pecan seedling rootstock treatments from high to low was as follows: ‘Posey’, ‘Peruque’, ‘Riverside’, ‘87MX5-1.7’, ‘VC1-68’, ‘Elliott’, ‘87MX1-2.2’, ‘San Felipe’, ‘Moore’, ‘Major’, ‘Giles’, and ‘Frutoso’.

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Rui Zhang, Fang-Ren Peng, Pan Yan, Fan Cao, Zhuang-Zhuang Liu, Dong-Liang Le, and Peng-Peng Tan

Root systems of pecan trees are usually dominated by a single taproot with few lateral roots, which are commonly thought to inhibit successful transplanting. This study aimed to evaluate early growth and root/shoot development of pecan seedlings in response to taproot pruning. Taproots of ‘Shaoxing’ seedling pecan trees were mildly (1/3 of the total length of the radicle removed) and severely (2/3 of the total length of the radicle removed) pruned at different seedling development stages shortly after germination. At the end of the first growing season, top growth was measured and then trees were uprooted so that root system regrowth could be evaluated. The results showed that root pruning had no impact on increases in stem height or stem diameter. However, pruning the taproot could stimulate primary growth in taproot branches. Root weight and the number of taproot branches per tree increased with decreasing taproot length. This study indicated that severe root pruning when three to five leaves had emerged resulted in root systems with more taproot branches and the greatest root dry weight after one growth season, which may increase survival and reduce transplanting shock.

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Rui Zhang, Fang-Ren Peng, Dong-Liang Le, Zhuang-Zhuang Liu, Hai-Yang He, You-Wang Liang, Peng-Peng Tan, Ming-Zhuo Hao, and Yong-Rong Li

Scion wood of ‘Caddo’ and ‘Desirable’ pecan (Carya illinoinensis) was grafted onto the epicotyl of 1-month-old, open-pollinated ‘Shaoxing’ pecan seedlings for evaluation as a grafting technique to reduce the time to produce grafted trees. The results showed that seedlings grafted with “base scions” had higher survival than those grafted with “terminal scions” for both ‘Caddo’ and ‘Desirable’. Also, grafting with paraffinic tape could achieve greater success rate than that with medical tape. The most ideal time to perform this grafting was late April in Nanjing, China, when pecan seedlings were about 35 days old. This study demonstrated that the technique yielded successful epicotyl grafting of >70%, and it could thus be applied in practice.

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Zhuang-Zhuang Liu, Tao Chen, Fang-Ren Peng, You-Wang Liang, Peng-Peng Tan, Zheng-Hai Mo, Fan Cao, Yang-Juan Shang, Rui Zhang, and Yong-Rong Li

Cytosine methylation plays important roles in regulating gene expression and modulating agronomic traits. In this study, the fluorescence-labeled methylation-sensitive amplified polymorphism (F-MSAP) technique was used to study variation in cytosine methylation among seven pecan (Carya illinoinensis) cultivars at four developmental stages. In addition, phenotypic variations in the leaves of these seven cultivars were investigated. Using eight primer sets, 22,796 bands and 950 sites were detected in the pecan cultivars at four stages. Variation in cytosine methylation was observed among the pecan cultivars, with total methylation levels ranging from 51.18% to 56.58% and polymorphism rates of 82.29%, 81.73%, 78.64%, and 79.09% being recorded at the four stages. Sufficiently accompanying the polymorphism data, significant differences in phenotypic traits were also observed among the pecan cultivars, suggesting that cytosine methylation may be an important factor underlying phenotypic variation. Hypermethylation was the dominant type of methylation among the four types observed, and full methylation occurred at higher levels than did hemimethylation in the pecan genomes. Cluster analysis and principal coordinate analysis (PCoA) identified Dice coefficients ranging from 0.698 to 0.778, with an average coefficient of 0.735, and the variance contribution rates of the previous three principal coordinates were 19.6%, 19.0%, and 18.2%, respectively. Among the seven pecan cultivars, four groups were clearly classified based on a Dice coefficient of 0.75 and the previous three principal coordinates. Tracing dynamic changes in methylation status across stages revealed that methylation patterns changed at a larger proportion of CCGG sites from the 30% of final fruit-size (30%-FFS) stage to the 70%-FFS stage, with general decreases in the total methylation level, the rate of polymorphism, and specific sites being observed in each cultivar. These results demonstrated that the F-MSAP technique is a powerful tool for quantitatively detecting cytosine methylation in pecan genomes and provide a new perspective for studying many important life processes in pecan.