种子老化影响玉米、大麦、小麦种质遗传完整性变化的研究
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摘要
研究种子老化对库存种质遗传完整性变化的影响，是种质保存中最亟待解决的问题，对于确保种质资源安全保存具有非常重要的现实意义。本研究主要以玉米、大麦和小麦种质作为研究对象，利用形态标记、细胞标记、生化标记以及DNA分子标记等方法，来探讨种子生活力下降对种质遗传完整性变化的影响，旨在为种质发芽率更新标准的制定提供科学的理论和实践依据。主要结果如下：
1、通过农艺性状和SSR分子标记技术对玉米地方品种条花糯不同发芽率水平的群体进行遗传分析。结果表明，同一份玉米种质材料，人工老化处理的低发芽率种质群体，其多态性条带百分率、等位基因数、有效等位基因数、基因多样性、Shannon指数等遗传参数数值，都较对照的高发芽率种质群体的遗传参数数值有所下降，表明同一玉米品种群体内，老化处理群体内的遗传多样性低于对照群体的遗传多样性，其群体内遗传变异出现下降。随着种子发芽率的下降，其出苗植株中芽鞘色为浅紫色植株样本的比率也随之下降，当发芽率降到32％时，其出苗植株中芽鞘色为浅紫色植株样本的比率降至0。该结果表明，当该份玉米种子发芽率降至32％时，其更新下一代的种质遗传组成与亲本种质的遗传组成可能发生了变化，即种质群体中已不含芽鞘色为浅紫的植株。另外，在不同发芽率出苗植株群体中，叶鞘色为浅紫色植株的比率有随发芽率下降而升高的趋势，而叶鞘色为白色植株的比率则表现相反，为下降的趋势。该结果也从另一方面证明，对于异质种质材料，低的发芽率更新标准是不利于确保种质遗传完整性的维持。
2、应用植物细胞染色体技术和醇溶蛋白酸性聚丙烯酰胺凝胶电泳技术对大麦地方品种普乃干木进行分析。实验结果表明，不同发芽率水平的老化种子，其染色体畸变率不存在显著差异。但随着种子生活力的下降，该种质群体醇溶蛋白的四种谱带类型的“biotype”频率也发生变化，当发芽率降到34％时，其中一种谱带类型“biotype”消失，即可理解为更新下一代的种质遗传组成可能发生了遗传上的漂移。同时也表明大麦醇溶蛋白分析方法应用于研究大麦种质遗传完整性变化是可行的。
3、对每一品种分别来自中期库(低生活力种子)和长期库(高生活力种子)的30份小麦种质进行农艺性状和生育期调查，发现在这两组之间，除了穗长有显著差异外，其它各项指标均无显著差异；对其中20份进行植物细胞染色体测定，结果表明中期库的小麦种质，其更新前种子根尖细胞平均染色体畸变率为4.41％，明显高于长期库种质的平均染色体畸变率(0.49％)。种子根尖染色体畸变率与发芽率呈负显著相关，但不管是贮藏于长期库或中期库，种子根尖染色体畸变率与贮藏年限均无显著相关。进一步对其中9份新繁殖子一代小麦种子进行分析，发现此时种子根尖细胞染色体畸变率已基本无差异。
The effect of seed aging on genetic integrity changes of stored seeds in genebank is an urgent task to be clarified for germplasm preservation. It is of great significance for a safe conservation of germplasm resources. Working on seeds of maize (ZeamaysL.) , barley (Hordeum vulgare L.) and wheat (Triticum aestivum L.) , the effect of seed viability decline on the alteration of genetic integrity was analyzed with morphological markers, cytological markers, biochemical markers and DNA molecular markers, so as to provide scientific and theoretical basis for making a germination-rate-based criteria for regenerating germplasm resources. Main results found were as the following.
1. Genetic alterations of maize landrace cv. Tiaohuanuo populations having different germination levels were analyzed by using agronomic traits and SSR markers. It was shown that change measured as the percentage of polymorphic bands, the number of alleles (na), effective number of alleles (ne) , Nei's gene diversity index (h) and Shannon's information index (I) and so on, were lower in artificially aged population with lower germination percentage than in CK population with higher germination percentage. This indicated that the genetic diversity was lower in aged population than in CK population. The proportion of plant with light-purple gemmule sheath decreased with the decline of germination percentage. When seed germination percentage declined to 32%, there was no light-purple plant. This result indicated that alternation of the genetic composition from light-purple plant to no light-purple plant could occur in regenerating population if the seed germination percentage declined to 32%. On the other h
and, the proportion of light-purple leaf sheath plant increased while the white leaf sheath plant decreased, when seed germination percentage declined. This result also indicated that a low germination percentage standard for regeneration is disadvantageous for maintaining the genetic integrity of heterogeneous accessions.
2. The hereditary characters of barley landrace cv. Punaiganmu were analyzed using botanical cell chromosomal and A-PAGE methods. The result showed that there was no significant difference in seed chromosomal aberration ratio in root tip cells between populations with different germination percentage. By the use of A-PAGE, four biotypes differing in the hordein spectra have been determined, biotype ratio varied with the decline of germination percentage. One "biotype" disappeared while seed germination percentage declined to 34%. The result indicated that the loss of seed viability and differential survival of the biotypes resulted in selection and genetic shifts in germplasm accessions, and also indicated that the A-PAGE could be used to detect genetic integrity change in barley accessions.
3. A same set of 30 wheat accessions from two sources, one set was from seeds stored in the National Long-term Genebank, and the other was from seeds stored in the mid-term Genebank. On growth
duration and other agronomic traits studies, no significant differences were found between two sets except on panicle length. Chromosomal aberration percentages in root tip cells of 20 wheat accessions which were stored in mid-term genebank and had lower viabilities were averaged as 4.41%, which was obviously higher than the value of 0.49% calculated from cells of wheat accessions which were stored in long-term genebank and had higher viability. Seed chromosomal aberration proportion in root tip cells was negatively correlated with seed germination percentage. There was no significant correlation between seed germination percentage and storage time for seeds no matter they were stored in the mid-term or long-term genebank. Studies on 9 wheat accessions newly regenerated also showed that there was no significant difference on the chromosomal aberration in root tip cells between seeds stored in mid-term genebank and long-term genebank.
1、通过农艺性状和SSR分子标记技术对玉米地方品种条花糯不同发芽率水平的群体进行遗传分析。结果表明，同一份玉米种质材料，人工老化处理的低发芽率种质群体，其多态性条带百分率、等位基因数、有效等位基因数、基因多样性、Shannon指数等遗传参数数值，都较对照的高发芽率种质群体的遗传参数数值有所下降，表明同一玉米品种群体内，老化处理群体内的遗传多样性低于对照群体的遗传多样性，其群体内遗传变异出现下降。随着种子发芽率的下降，其出苗植株中芽鞘色为浅紫色植株样本的比率也随之下降，当发芽率降到32％时，其出苗植株中芽鞘色为浅紫色植株样本的比率降至0。该结果表明，当该份玉米种子发芽率降至32％时，其更新下一代的种质遗传组成与亲本种质的遗传组成可能发生了变化，即种质群体中已不含芽鞘色为浅紫的植株。另外，在不同发芽率出苗植株群体中，叶鞘色为浅紫色植株的比率有随发芽率下降而升高的趋势，而叶鞘色为白色植株的比率则表现相反，为下降的趋势。该结果也从另一方面证明，对于异质种质材料，低的发芽率更新标准是不利于确保种质遗传完整性的维持。
2、应用植物细胞染色体技术和醇溶蛋白酸性聚丙烯酰胺凝胶电泳技术对大麦地方品种普乃干木进行分析。实验结果表明，不同发芽率水平的老化种子，其染色体畸变率不存在显著差异。但随着种子生活力的下降，该种质群体醇溶蛋白的四种谱带类型的“biotype”频率也发生变化，当发芽率降到34％时，其中一种谱带类型“biotype”消失，即可理解为更新下一代的种质遗传组成可能发生了遗传上的漂移。同时也表明大麦醇溶蛋白分析方法应用于研究大麦种质遗传完整性变化是可行的。
3、对每一品种分别来自中期库(低生活力种子)和长期库(高生活力种子)的30份小麦种质进行农艺性状和生育期调查，发现在这两组之间，除了穗长有显著差异外，其它各项指标均无显著差异；对其中20份进行植物细胞染色体测定，结果表明中期库的小麦种质，其更新前种子根尖细胞平均染色体畸变率为4.41％，明显高于长期库种质的平均染色体畸变率(0.49％)。种子根尖染色体畸变率与发芽率呈负显著相关，但不管是贮藏于长期库或中期库，种子根尖染色体畸变率与贮藏年限均无显著相关。进一步对其中9份新繁殖子一代小麦种子进行分析，发现此时种子根尖细胞染色体畸变率已基本无差异。
The effect of seed aging on genetic integrity changes of stored seeds in genebank is an urgent task to be clarified for germplasm preservation. It is of great significance for a safe conservation of germplasm resources. Working on seeds of maize (ZeamaysL.) , barley (Hordeum vulgare L.) and wheat (Triticum aestivum L.) , the effect of seed viability decline on the alteration of genetic integrity was analyzed with morphological markers, cytological markers, biochemical markers and DNA molecular markers, so as to provide scientific and theoretical basis for making a germination-rate-based criteria for regenerating germplasm resources. Main results found were as the following.
1. Genetic alterations of maize landrace cv. Tiaohuanuo populations having different germination levels were analyzed by using agronomic traits and SSR markers. It was shown that change measured as the percentage of polymorphic bands, the number of alleles (na), effective number of alleles (ne) , Nei's gene diversity index (h) and Shannon's information index (I) and so on, were lower in artificially aged population with lower germination percentage than in CK population with higher germination percentage. This indicated that the genetic diversity was lower in aged population than in CK population. The proportion of plant with light-purple gemmule sheath decreased with the decline of germination percentage. When seed germination percentage declined to 32%, there was no light-purple plant. This result indicated that alternation of the genetic composition from light-purple plant to no light-purple plant could occur in regenerating population if the seed germination percentage declined to 32%. On the other h
and, the proportion of light-purple leaf sheath plant increased while the white leaf sheath plant decreased, when seed germination percentage declined. This result also indicated that a low germination percentage standard for regeneration is disadvantageous for maintaining the genetic integrity of heterogeneous accessions.
2. The hereditary characters of barley landrace cv. Punaiganmu were analyzed using botanical cell chromosomal and A-PAGE methods. The result showed that there was no significant difference in seed chromosomal aberration ratio in root tip cells between populations with different germination percentage. By the use of A-PAGE, four biotypes differing in the hordein spectra have been determined, biotype ratio varied with the decline of germination percentage. One "biotype" disappeared while seed germination percentage declined to 34%. The result indicated that the loss of seed viability and differential survival of the biotypes resulted in selection and genetic shifts in germplasm accessions, and also indicated that the A-PAGE could be used to detect genetic integrity change in barley accessions.
3. A same set of 30 wheat accessions from two sources, one set was from seeds stored in the National Long-term Genebank, and the other was from seeds stored in the mid-term Genebank. On growth
duration and other agronomic traits studies, no significant differences were found between two sets except on panicle length. Chromosomal aberration percentages in root tip cells of 20 wheat accessions which were stored in mid-term genebank and had lower viabilities were averaged as 4.41%, which was obviously higher than the value of 0.49% calculated from cells of wheat accessions which were stored in long-term genebank and had higher viability. Seed chromosomal aberration proportion in root tip cells was negatively correlated with seed germination percentage. There was no significant correlation between seed germination percentage and storage time for seeds no matter they were stored in the mid-term or long-term genebank. Studies on 9 wheat accessions newly regenerated also showed that there was no significant difference on the chromosomal aberration in root tip cells between seeds stored in mid-term genebank and long-term genebank.
引文
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