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端粒是位于真核生物染色体末端的核蛋白结构,它由端粒DNA重复序列、组蛋白八聚体和蛋白端粒帽子共同形成[1-2]。大多数陆生植物的端粒DNA是拟南芥式的7核苷酸重复(TTTAGGG)n[3],但是也有人类式重复等很多例外情况存在(表 1)[2]。端粒能够保护线性染色体不被内源核酸酶损伤,避免线性染色体被误识为未配对的染色体断片[2];端粒与细胞分裂、细胞衰老及寿命有着密切的联系[4]。在人类[5-6]和澳大利亚海狮等动物体细胞中[7-9],端粒随DNA复制和细胞分裂次数的增加而缩短。当端粒短到一定程度,细胞进入衰老期[10]。因此,人类和动物体细胞的端粒缩短被认为是老化的指标。虽然在大麦[11]和人参[9]等植物中也检测到年龄和端粒长度呈显著相关[4],但是植物中端粒长度和年龄的关系却远比动物复杂得多。随着植物年龄的增长和发育的进行,有些物种有些部位的端粒长度呈延长趋势、有些缩短、有些基本不变,甚至有些呈现周期性变化(表 2)。总之,不同植物物种、同一物种不同基因型、同一物种不同组织、同一物种不同分化程度的相同组织、不同环境及生理条件下的相同部位,端粒长度都可能存在差异[3, 9]。尽管如此,针对有些多年生植物物种的研究发现,端粒长度可以用来预测植株的年龄。例如,2015年Liang等人建立了人参年龄和端粒长度相关的数学模型[9];2015年刘頔等人建立了油松端粒长度和树龄的相关关系模型[4]。植物体的端粒调控存在复杂的机制:端粒酶可以在一定程度上维持或者延长端粒DNA的长度[22];在缺乏端粒酶的情况下,端粒可被“依赖于重组的替代机制”所延长[23];新近研究发现多种表观遗传机制参与了端粒稳定性[24-25]。可见关于植物端粒长度的维持机制,仍有很多内容有待研究。
表 1 目前发现的陆生植物端粒DNA多样性(非拟南芥式重复)
Table 1. The current knowledge on telomere DNA diversity in land plants (The non-Arabidopsis-type of telomeric repeats)
物种
Species端粒DNA重复单元
Repeat units of telomeric DNA蒜(Allium sativum) CTCGGTTATGGG 洋葱(Allium cepa) CTCGGTTATGGG 琉璃苣(Borago officinalis) TTAGGG 鸢尾属(Iris) TTAGGG 矛花属(Doryanthes) TTTAGGG 蝴蝶兰属(Phalaenopsis) TTTAGGG 玉米(Zea mays) TTTAGGG 螺旋狸藻属一种(Genlisea hispidula) TTCAGG 螺旋狸藻属一种(Genlisea nigrocaulis) TTTCAGG 番茄(Lycopersicon esculentum) TTTAGGG 马铃薯(Solanum tuberosum) TTTAGGG 烟草(Nicotiana tabacum) TTTAGGG 夜香树属(Cestrum) TTTTTTAGGG 南芥属(Arabis) TTTAGGG 松属(Pinus) TTTAGGG 卷柏属(Selaginella) TTTAGGG 苔藓(Moss) TTTAGGG 表 2 目前发现的植物端粒长度随年龄、发育和细胞分裂的变化情况
Table 2. The current knowledge on telomere length changes in plants during aging, development and cell division
变化趋势分组
Grouping according to changing trend植株、器官或组织
Plant, organ or tissue不变组 苹果树[12] 草莓植株[12] 1200年生苏格兰松形 成层、芽和针叶[13] 番茄叶[14] 女娄菜植株[15] 烟草愈伤组织[16] 白桦树[3] 延长组 银杏叶、愈伤组织[17] 人参根顶端[9] 大麦愈伤培养后期[18] 油松当年生枝条针叶[4] 以女娄菜叶为外植体培 养3个月的愈伤[19] 缩短组 200年以上苏格兰松茎 形成层[13] 大麦愈伤培养初期[18] 大麦植株[18] 秋季银杏叶[20] 白桦茎形成层(越往树顶越短)[3] 白桦组织培养过程中的材料[3] 循环变化组 刺果松针叶[21] 植物离体快速繁殖,即植物组织培养,是植物无性繁殖的典范。它是利用植物细胞的全能性,在无菌条件下,对离体的植物器官、组织、细胞及原生质体进行培养,以获得完整植株的技术。尽管体细胞无性系变异广泛存在[26-29], 较之有性繁殖技术,植物组织培养技术具备繁殖速度快,繁殖植株能在更大程度上保持母株的优良性状等诸多优点[26]。在林业领域,植物离体快速繁殖具备更加重要的意义。因为林木通常具备生长周期长、子代性状分离显著等特点。一旦发现优良单株,比如杂种优势成年优株,利用离体快速繁殖可以实现短期繁殖大量优良植株的目的。值得注意的是,2014年Aronen和Ryynänen研究发现随着白桦愈伤组织离体培养时间的延长,其端粒长度呈逐渐缩短的趋势[3]。又因为端粒长度也可以用于预测植物细胞的分裂能力和植株的年龄[4, 9, 30],所以以上发现引发了我们对于无性繁殖植株生理年龄的思考,本文将围绕端粒长度这条主线探讨无性繁殖植株的生理年龄。
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目前关于植物端粒长度和年龄相关的研究报道仍然很有限[9, 36]。很多植物物种,尤其是多年生林木的无性繁殖植株的端粒长度及寿命有待研究。外植体的端粒长度是否可以用于预测再生能力,愈伤组织的端粒长度是否可以预测再分化能力,何时诱导愈伤组织再分化更合适?什么样的愈伤细胞才能成为再分化的感受态细胞?嫁接诱导老树复幼是否与端粒长度的变化有关?组培再生植株的幼龄化表型是否与端粒的长度有关?这些都是亟需研究和解决的科学问题。例如,仅仅对于用叶片作为外植体的离体快速繁殖,就有很多内容是值得研究的(表 3)。尤其对于叶片端粒长度与植株其它部位存在明显差异的物种。我们坚信随着高通量测序等实验技术手段的不断进步,这些问题在不久的将来都将被揭晓。
表 3 不同材料端粒长度比较可能揭示的内容
Table 3. The substance which might be revealed by comparing telomere length among different materials
用于比较端粒长度的材料
Materials used for comparing length of telomere拟揭示的内容
Substance to be revealed细胞全能性叶片vs. 细胞无全能性叶片 全能性与端粒长度的相关性 外植体叶片vs. 叶片直接发生植株 端粒长度与直接发生的关系 外植体叶片vs. 叶片产生愈伤组织 脱分化及愈伤增殖与端粒长度的相关性 叶片产生无分化能力愈伤vs.有分化能力愈伤 端粒长度与分化能力的相关性 叶片产生玻璃化愈伤vs.正常没玻璃化愈伤 愈伤玻璃化与端粒长度的相关性 叶片产生愈伤组织vs. 叶片间接发生植株 再分化与端粒长度的相关性 外植体叶片vs. 叶片间接发生植株 外植体对间接植株端粒长度的影响 叶片直接发生植株vs. 叶片间接发生植株 愈伤阶段对端粒长度的影响 同等发育程度实生苗vs. 叶片间接发生植株 直接发生植株的生理年龄和预后 同等发育程度实生苗vs. 叶片直接发生植株 间接发生植株的生理年龄和预后
无性繁殖植株的生理年龄——由端粒长度引发的思考
The Physiological Age of Asexual Plants——Thinking Arise from Telomere Length
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摘要: 植物细胞中端粒长度随年龄的变化远比动物复杂,可分为不变组、延长组、缩短组和循环变化组。尽管如此,端粒长度亦可用于预测植物年龄。在植物组织培养过程中,端粒长度也会变化。故无性繁殖植株生理年龄值得思考和研究。本文基于端粒长度探讨了无性繁殖植株生理年龄。认为生理年龄较老供体可能会无性繁殖出"小老头"无性系,端粒长度可能成为预测植物材料无性繁殖能力和愈伤组织再分化能力的有效新指标,老树复幼和端粒长度的关系是一个很好的研究课题。此外,提出了植物愈伤组织需要足够的时间来抹除外植体"痕迹"的新观点。关于植物无性繁殖过程中的端粒长度,有很多问题亟待研究和探索。Abstract: The change of telomere length during aging of plant cells has been found to be much more complicated than that in animals. It can be divided into the invariant group, the prolongation group, the shortening group and the circle change group. Nevertheless, the age of plants can also be predicted based on telomere length. Moreover, the telomere length can change during plant tissue culture. Thus, the physiological age of asexual plants is worth to be studied. In the article, the physiological age of asexual plants is discussed based on telomere length. It is concluded that the older donor might produce clone with small body but old physiological age, telomere length might be a novel reliable indicator for assessment the ability of both asexual propagation and callus redifferentiation of plants, the relationship between old-tree reinvigoration and telomere length might be a good research project. Furthermore, it is proposed that plant calli need enough time to erase the traces of the explants. Many problems about the telomere length during asexual propagation of plants are urgent to be researched and explored.
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Key words:
- telomere
- / in vitro rapid propagation
- / cutting
- / grafting
- / physiological age
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表 1 目前发现的陆生植物端粒DNA多样性(非拟南芥式重复)
Table 1. The current knowledge on telomere DNA diversity in land plants (The non-Arabidopsis-type of telomeric repeats)
物种
Species端粒DNA重复单元
Repeat units of telomeric DNA蒜(Allium sativum) CTCGGTTATGGG 洋葱(Allium cepa) CTCGGTTATGGG 琉璃苣(Borago officinalis) TTAGGG 鸢尾属(Iris) TTAGGG 矛花属(Doryanthes) TTTAGGG 蝴蝶兰属(Phalaenopsis) TTTAGGG 玉米(Zea mays) TTTAGGG 螺旋狸藻属一种(Genlisea hispidula) TTCAGG 螺旋狸藻属一种(Genlisea nigrocaulis) TTTCAGG 番茄(Lycopersicon esculentum) TTTAGGG 马铃薯(Solanum tuberosum) TTTAGGG 烟草(Nicotiana tabacum) TTTAGGG 夜香树属(Cestrum) TTTTTTAGGG 南芥属(Arabis) TTTAGGG 松属(Pinus) TTTAGGG 卷柏属(Selaginella) TTTAGGG 苔藓(Moss) TTTAGGG 表 2 目前发现的植物端粒长度随年龄、发育和细胞分裂的变化情况
Table 2. The current knowledge on telomere length changes in plants during aging, development and cell division
变化趋势分组
Grouping according to changing trend植株、器官或组织
Plant, organ or tissue不变组 苹果树[12] 草莓植株[12] 1200年生苏格兰松形 成层、芽和针叶[13] 番茄叶[14] 女娄菜植株[15] 烟草愈伤组织[16] 白桦树[3] 延长组 银杏叶、愈伤组织[17] 人参根顶端[9] 大麦愈伤培养后期[18] 油松当年生枝条针叶[4] 以女娄菜叶为外植体培 养3个月的愈伤[19] 缩短组 200年以上苏格兰松茎 形成层[13] 大麦愈伤培养初期[18] 大麦植株[18] 秋季银杏叶[20] 白桦茎形成层(越往树顶越短)[3] 白桦组织培养过程中的材料[3] 循环变化组 刺果松针叶[21] 表 3 不同材料端粒长度比较可能揭示的内容
Table 3. The substance which might be revealed by comparing telomere length among different materials
用于比较端粒长度的材料
Materials used for comparing length of telomere拟揭示的内容
Substance to be revealed细胞全能性叶片vs. 细胞无全能性叶片 全能性与端粒长度的相关性 外植体叶片vs. 叶片直接发生植株 端粒长度与直接发生的关系 外植体叶片vs. 叶片产生愈伤组织 脱分化及愈伤增殖与端粒长度的相关性 叶片产生无分化能力愈伤vs.有分化能力愈伤 端粒长度与分化能力的相关性 叶片产生玻璃化愈伤vs.正常没玻璃化愈伤 愈伤玻璃化与端粒长度的相关性 叶片产生愈伤组织vs. 叶片间接发生植株 再分化与端粒长度的相关性 外植体叶片vs. 叶片间接发生植株 外植体对间接植株端粒长度的影响 叶片直接发生植株vs. 叶片间接发生植株 愈伤阶段对端粒长度的影响 同等发育程度实生苗vs. 叶片间接发生植株 直接发生植株的生理年龄和预后 同等发育程度实生苗vs. 叶片直接发生植株 间接发生植株的生理年龄和预后 -
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