| 王春梅,沈珊,李景军,张秋明,曹尚银.枫杨砧木与核桃嫁接接合部愈合过程的解剖学研究[J].中国南方果树,2022,51(1): |
| 枫杨砧木与核桃嫁接接合部愈合过程的解剖学研究 |
| Anatomical study on the healing process of the grafting joint between Pterocarya stenoptera rootstock and Juglans regia |
| 投稿时间：2021-10-11 修订日期：2021-10-12 |
| DOI： |
| 中文关键词: 枫杨 核桃 嫁接 愈伤组织 显微结构 |
| 英文关键词:Pterocarya stenoptera, Walnut, grafting, callus,Microstructure. |
| 基金项目:中国农业部果树育种技术重点实验室项目（NYB201603-2）;湖南省教育厅科学研究优秀青年项目(20B593)；中国农学会教育教学类科研课题(PCE2032);岳阳职业技术学院科研团队项目(洞庭湖区域乡村振兴战略实施研究)；岳阳市社会科学研究课题(2020Y76)；岳阳职业技术学院院级课题(yz20-08)。 |
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| 中文摘要: |
| [目的]揭示核桃/枫杨嫁接组合嫁接愈合过程组织细胞学特征。[方法]以2年生湖南本地枫杨实生苗为砧木,以“中核短枝”核桃品种的1年生枝条为接穗,采用改良的方块芽接技术嫁接,取其嫁接接合部位组织,运用解剖学方法,通过光学显微镜及透射电镜进行嫁接愈合过程的组织细胞学观察。[结果]通过嫁接口愈合过程的显微结构解剖学观察,发现其愈合过程经过了隔离层形成期、愈伤组织形成期、形成层环形成期、维管组织分化形成期四个时期；通过超微结构观察发现,嫁接初期出现嫁接切口面附近细胞线粒体、淀粉粒数量增加,出现转运物质的壁旁体、多泡体等,细胞质呈现油滴现象等系列愈伤反应。[结论]嫁接后的前20d左右形成愈伤组织桥、形成层环、维管束桥,是嫁接成活的关键时期。形成层不是产生愈伤组织的唯一来源,皮层也可形成愈伤组织。嫁接初期隔离层两边的细胞会出现细胞器变化,内膜系统重组,物质转移等系列愈伤反应。 |
| 英文摘要: |
| [Objective] to reveal the Histocytological characteristics of walnut / Pterocarya stenoptera grafting combination during grafting healing[ [method] the 2-year-old seedlings of Pterocarya stenoptera in Hunan Province were used as rootstock, and the 1-year-old branches of "Zhonghe short branch" walnut variety were used as scions. The improved square budding grafting method was adopted. The grafting time was June 20. The samples were collected at 3, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 30 and 45 days after grafting. The length of phloem and callus is 5-6 mm. The samples were fixed with FAA (containing 70% ethanol) and stored at 4 ℃ for slicing. Fixed with 2.5% glutaraldehyde for 2 to 4 hours, washed with phosphoric acid buffer for 3 times, 1% osmic acid for 4 to 6 hours, washed with buffer for 3 times, washed for 20 minutes each time, dehydrated with 30%, 50%, 70%, 80%, 95% pure acetone, dehydrated with alcohol step by step at room temperature, each stage was used for 30 minutes, acetone was changed for 3 times, and embedded with EPON 812 epoxy resin. It was made by leciarm2245 rotary microtome, 10 μm 5% periodate to remove the resin and osmium, toluidine blue staining, Leica dm5000 light microscope observation and photography. Ultrathin sections with thickness of 50-70nm were made by ultrathin sectioning machine, stained with uranium acetate and lead citrate, and observed and photographed by h-7800 transmission electron microscope.[ Results] through the longitudinal optical microscope, the structures of walnut scion and rootstock Pterocarya stenoptera were pericarp, cortex, phloem, cambium and xylem. They also have developed phloem and a large number of phloem fibers. Among them, the phloem fiber of rootstock has single row and double row, and the scion is mostly single row, with high degree of similarity in tissue structure. Through the observation of microstructural anatomy, it was found that the healing process of the interface went through four stages: isolation layer formation stage, callus formation stage, cambium ring formation stage and vascular tissue differentiation stage. Three days after grafting, isolation layer was formed between walnut scion and maple poplar rootstock; At 6th day after grafting, soft callus parenchyma cells were formed on both sides of the isolation layer between rootstock and scion; Callus bridge was formed 10 days after grafting; 20 days after grafting, with the continuous proliferation of callus, tubular and sieve molecules were differentiated in some areas of callus, which connected and penetrated each other, and then formed vascular bundle bridge connecting rootstock and spike; After 30 days, the new vascular cambium formed secondary phloem outward, contained new sieve tubes, formed xylem inward,
formed secondary xylem inward, and produced new vessels. Therefore, the formation of callus bridge, cambium ring and vascular bundle bridge in the 20 days after grafting is the key period for grafting survival; At the same time, we also observed the following phenomena: cortex parenchyma cells can also form a large number of calli, cambium is not the only source of callus , cortex parenchyma cells form a large number of calli, scion and rootstock callus, parenchyma cells fusion form, callus bridge form, layer ring form, Vascular bundle bridge and new vascular tissue morphology. In the early stage of grafting, there were a series of callus reactions, such as the increase of the number of mitochondria and starch grains near the surface of grafting incision, the appearance of paracellular bodies and polyvesicles transporting substances, and the appearance of oil drops in the cytoplasm. After 3-14 days of grafting, the cells on the surface of grafting incision were damaged due to cutting injury, the cambium cells were stimulated by trauma, the protoplasts of cells were degraded or disappeared slowly, some cell walls were broken, and the cytoplasm was squeezed to form isolation layer. After the cell wall of adjacent parenchyma cells ruptured, the disintegrated flocculent protoplasts fused with each other. In the severely injured cells, the central large vacuoles disappeared and many small vacuoles appeared, and the cytoplasm was cotton wadding. In phloem and xylem, the plasmodesmata between parenchyma cells are obvious, and they often form in groups at the thinner cell wall. In the early stage of grafting, the parenchyma cells of calli on both sides of cambium secreted pectin outside the cell wall, which was bead like and sandwiched in the middle of the adjacent cell wall. With the accumulation of pectin, the parenchyma cells of calli increased significantly. 30-45 days after grafting, the grafting was successful, and the pectin gradually dispersed and disappeared in the cell wall complex, Plasmodesmata were found in the thin cell wall. There were many small vacuoles, abundant protein and mitochondria in the cambium zone cells, and the cytoplasm was flocculent, which still showed obvious callus reaction [Conclusion] callus bridge, cambium ring and vascular bundle bridge were formed about 20 days after grafting, which was the key period for grafting survival. Cambium is not the only source of callus, and cortex can also form callus. At the early stage of grafting, a series of callus reactions, such as organelle changes, reorganization of endomembrane system and material transfer, occurred in the cells on both sides of the isolation layer. |
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