红松种子发育过程中内源激素含量的动态变化

Content Dynamics of Endogenous Hormones in Different Seed Developmental Stages of Korean Pine

【目的】生长素、细胞分裂素、赤霉素、脱落酸等植物内源激素在促进种子胚胎发育进程的演变过程中有重要作用。本研究通过分析红松胚胎发育过程中4个重要阶段雌配子体(含合子胚)的IAA,ZR,GAs,ABA 4种内源激素的动态变化,拟为解释红松胚胎发育机制提供证据,为体胚诱导过程中添加外源激素及红松种子园种子发育过程的外源激素调控提供参考。【方法】以处于原胚期、裂生多胚期、柱状胚期、子叶胚前期的红松雌配子体作为研究材料,监测红松胚胎发育形态演变过程,同时采用酶联免疫法(ELISA)测定2个无性系不同发育时期红松种子内源激素生长素(IAA)、赤霉素(GAs)、玉米素核苷(ZR)、脱落酸(ABA)含量的变化。【结果】2个无性系的内源激素含量变化趋势相似,红松胚胎发育4个阶段IAA和ABA含量均维持在较高的水平,GAs含量全程较低。其中ZR含量在胚胎发育全程呈现下降的变化趋势,原胚期ZR含量最高,无性系057和059中平均含量分别为12.901和13.622 4 ng·g^(-1)FW;IAA的含量先缓慢上升随后急剧上升,柱状胚期至子叶胚期含量急剧下降,柱状胚期含量达到整个发育过程中的最高值,无性系057和059平均含量分别为101.923 0和107.256 4 ng·g^(-1)FW;GAs含量呈现先降低后增加的变化趋势,原胚期至裂生多胚期的GAs含量呈现递减的变化趋势,裂生多胚期含量最低,无性系057和059含量分别为4.823 5和5.588 8 ng·g^(-1)FW,发育至柱状胚期含量呈现上升趋势,并且该趋势一直持续至子叶胚前期,子叶胚前期与原胚期GAs含量差异不显著;ABA含量呈现先上升后递减的变化趋势,具体表现为在原胚期含量为4个发育期的最低,无性系057和059含量分别为88.497 4和94.847 3 ng·g^(-1)FW,随后一直保持上升的趋势直至发育至裂生多胚期达到最高水平,无性系057和059含量分别为133.200 1和139.983 9 ng·g^(-1)FW,在随后的发育过程中ABA含量逐渐降低,2个无性系的ABA含量分别接近各自原胚期的水平;GAs+IAA+ZT/ABA的值呈现先增加后降低趋势,柱状胚期达到最大值。【结论】红松种子发育过程中内源激素的含量变化随着胚胎形态发育而发生动态变化,各呈现不同的变化格局;IAA,ABA,ZR在红松胚胎发育过程中起重要调节作用,而GAs对发育过程没有明显的作用。

【Objective】The plant endogenous hormones,including auxin,cytokinin,gibberellin,and abscisic acid,play an important role in regulating the embryonic development of seeds.The dynamics of IAA,ZR,GAs,and ABA were studied in order to reveal the mechanism of embryonic development of Korean pine（ Pinus koraiensis） seed,as well as to provide evidence for adjusting the addition of exogenous hormones during somatic embryogenesis and / or seed development of in Korean pine seed orchard.【Method 】The morphological changes of the megagametophytes of Korean pine were observed during different developmental stages,including pro-embryo,cleavage polyembryony,columnar embryo,and pre-cotyledonary embryo,and the concentration of IAA,GAs,ZR,and ABA of two Korean pine clones（057 and 059,respectively） during the four embryonic development stages was analyzed using enzyme-linked immunosorbent assay（ELISA）.【Results】The concentration of the endogenous hormones at the four stages of embryonic development wassimilar in the two clones.The content of IAA and ABA during the four stages of embryonic development of Korean pine seed was maintained at a high level.In contrast,the content of GAs remained relatively low level during the whole processes.The concentration of ZR showed a downward tendency with the development of the embryo,with the peak values being found in the pro-embryo megagametophytes： 12.901 ng·g^（-1） FW in clone 057 and 13.622 4 ng·g^（-1）FW in clone 059.The IAA content increased slowly at first and then sharply at the stage of columnar,and then dramatically declined at the stage of pre-cotyledonary embryo.The IAA content in the columnar embryo was highest：101.923 0 and107.256 4 ng·g^（-1） FW for clone 057 and 059,respectively.The GAs content first decreased and then increased.In detail,the GAs content decreased from pro-embryo to cleavage polyembryony,with a lowest concentration of 4.823 5 and 5.588 8 ng·g^（-1）FW at the cleavage polyembryony stage for clone 057 and 059,respectively.After that,its GAs content started to increase again until the columnar embryo and all the way to pre-cotyledonary embryo stage,however,no significant difference in the GAs content was found between pro-embryo and pre-cotyledonary embryo stage.The ABA content showed a decreasing trend after the first rise.Specifically,the ABA content was lowest in pro-embryo stage and the ABA content of clone 057 and 059 was 88.497 4 and 94.847 3 ng·g^（-1）FW,respectively.Afterwards,a upward trend had been maintained until to the cleavage polyembryony stage,with the content reaching to the peak value of 133.200 1and 139.983 9 ng·g^（-1）FW for clone 057 and 059,respectively.At the following development stages the ABA content kept decreasing and returned to the level close to pro-embryo stage.The ratio of GAs＋IAA＋ZT to ABA increased at first,reached the maximum at the stage of the columnar embryo,and then decreased.【Conclusion】The content of endogenous hormones of Korean pine seeds showed a dynamic changes with the development of embryo morphology,and each hormone displayed different change pattern.In summary,IAA,ABA and ZR played an important regulatory role in embryonic development of Korean pine seeds,whereas GAs had no significant effects.