外源物质对不同倍性白花泡桐体外植株再生及幼苗蛋白质变化影响研究
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摘要
泡桐是一种深受我国广大劳动人民喜爱的速生用材和绿化树种,但泡桐生产中一直存在的丛枝病发生严重和低干大冠等问题,严重影响了人们种植泡桐的积极性。由于泡桐愈伤组织诱导时间短并且处于相同分裂周期的愈伤组织较多,有利于生理生化的研究;本试验利用DNA甲基化转移酶抑制剂(5-Aza)和组蛋白去乙酰化转移酶抑制剂(苯丁酸、Chaps)对二倍体和四倍体白花泡桐体外植株再生的影响以及幼苗形态蛋白的变化,进而探讨5-Aza、苯丁酸和Chaps对泡桐叶片芽诱导、幼苗形态变化、蛋白质表达结果,最终了解5-Aza、苯丁酸和Chaps对泡桐生长发育的影响。主要结果如下:
1外源物质抑制泡桐体外植株再生5-Aza、Chaps、苯丁酸对二倍体和四倍体白花泡桐的叶片芽诱导具有抑制作用,并且随着5-Aza、Chaps、苯丁酸浓度的升高抑制作用增强;5-Aza、Chaps、苯丁酸对二倍体和四倍体白花泡桐的根诱导具有抑制作用,随着5-Aza、Chaps、苯丁酸浓度的升高抑制作用增强。外源物质对二倍体白花泡桐的体外植株再生抑制作用大于四倍体白花泡桐体外植株再生。
2外源物质对幼苗形态变化产生明显影响用5-Aza、Chaps、苯丁酸处理二倍体和四倍体白花泡桐幼苗形态变化存在一定的差异,经5-Aza处理的泡桐出现了芽丛生和植株矮化,苯丁酸处理的泡桐均出现了根部有大量愈伤组织,伴随5-Aza、苯丁酸浓度的增加,幼苗的症状越明显。经Chaps处理的二倍体和四倍体白花泡桐泡桐幼苗形态变化不明显,随着外源物质浓度的增大生长抑制作用增强。
3外源物质对幼苗叶片蛋白质的变化产生了影响经5-Aza、Chaps、苯丁酸处理二倍体和四倍体白花泡桐幼苗进行蛋白质双向电泳发现,等电点、分子量、蛋白质含量在不同外源物质、同种外源物质不同浓度处理的幼苗中存在着差异。经5-Aza处理的二倍体白花泡桐幼苗,蛋白质PI 6.6、22KD(a), PI 6.5、21.8KD(b), PI 5.4、14.5 KD(k)增强表达,蛋白PI 5.2、17.3 KD(e) ,PI 5.2、16.7KD(f)减弱表达;而经5-Aza处理四倍体白花泡桐叶片蛋白质PI 6.6、22KD(a), PI 6.5、21.8KD(b)逐渐渐弱;蛋白质PI 5.2、17.3 KD(e)和PI 5.8、PI 5.2、16.7KD(f)逐渐减弱直至消失。经苯丁酸处理二倍体白花泡桐叶片出现特异蛋白质5.2、17.3 KD(e),5.8、16.5 KD(o);经苯丁酸处理四倍体白花泡桐叶片蛋白质PI 6.6、22KD(a),PI 6.5、21.8KD(b),PI 5.2、16.7KD(f)是差异蛋白质。经Chaps处理白花泡桐叶片蛋白质可以发现,随着Chaps浓度增加蛋白质PI 6.4、20.1 KD(p),PI 6.56、19.8 KD(q)逐渐增强,并且出现了差异蛋白质PI 5.2、17.3 KD(e);经chaps处理四倍体白花泡桐叶片蛋白质双向电泳蛋白PI 6.6、22KD(a), PI 6.5、21.8KD(b)先减弱后增强,PI 5.2、16.7KD(f)一直减小。
The pawlownia is one of our country important fast-growing timber wood tree seeds, is improving our country ecological environment and raises the people’s living standards.,but People are seriously affected the enthusiasm of the cultivation of Paulownia. Paulownia callus induction is as a result of a short time and in the same cycle of callus to split more conducive to study physiology and biochemistry;
To study the transcription of DNA methylation inhibitors (5-Aza) and histone- deacetvlase (4-Phenylbutyric acid, Chaps) on the Growth of Paulownia, the experiment with using two-dimensional gel electrophoresis analysis of proteins after 5-Aza, 4-Phenylbutyric acid, Chaps treatment on P. fortunei diploid and tetraploid P. fortunei differences in gene expression product; study of 5-Aza, 4-Phenylbutyric acid ,Chaps, in cell dedifferentiation process role, and then explore the DNA methylation inhibitor (5-Aza) and histone-deacetvlase acetyltransferase inhibitor (4-Phenylbutyric acid, Chaps) of Paulownia leaf bud induction, seedling morphology, protein expression results, Learn eventually 5-Aza, 4-Phenylbutyric acid, Chaps on the growth and development of Paulownia. The main findings are as follows:
1 Inhibition of exogenous substances in vitro plant regeneration of Paulownia 5-Aza, Chaps, 4-Phenylbutyric acid, treatment P. fortunei diploid and tetraploid and tetraploid has in vitro inhibitory effect of plant regeneration, and with 5 -Aza, Chaps, 4-Phenylbutyric acid, enhanced inhibitory concentration increased.
2 Exogenous substances on seedling have effect on morphological changes With 5-Aza, Chaps, 4-Phenylbutyric acid reatment on P. fortunei diploid and tetraploid and tetraploid morphology differences must exist, with 5-Aza deal with the Paulownia appeared numerous buds and plant dwarfing, 4-Phenylbutyric acid treatment of both the roots of Paulownia have substantial callus, with 5-Aza, Chaps, 4-Phenylbutyric acid concentration increased, seedlings, the more obvious symptoms. With Chaps paulownia seedlings of three treatment patterns did not change significantly.
3 Exogenous substances on diploid and tetraploid P. fortunei seedlings the leaves have effect on proteins Leaves of seelding by 5-Aza, Chaps, 4-Phenylbutyric acid to deal with found in two-dimensional gel electrophoresis protein isoelectric point, molecular weight, protein content, the same species of different concentrations of exogenous substances to deal with the difference between the presence of seedlings in the treatment of seedlings differences exist. After dealing with 5-Aza diploid P. fortunei seedlings, protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b), PI 5.4,14.5 KD (k) to enhance the expression of protein PI 5.2,17.3 KD (e ), PI 5.2, 16.7KD (f) reduced expression; while dealing with 5-Aza tetraploid P. fortunei leaf protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b) a gradual diminuendo; protein PI 5.2, 17.3 KD (e) and PI 5.8, PI 5.2, 16.7KD (f) until the disappearance of warning. Paulownia leaves handling by the 4-Phenylbutyric acid appear diploid-specific protein 5.2,17.3 KD (e), 5.8,16.5 KD (o); by 4-Phenylbutyric acid handling protein tetraploid P. fortunei leaves PI 6.6, 22KD (a), PI 6.5, 21.8KD (b), PI 5.2, 16.7KD (f) is the difference in protein. Chaps by P. fortunei leaf protein processing can be found, with the increase in protein. Chaps concentration PI 6.4,20.1 KD (p), PI 6.56,19.8 KD (q) gradually increased, and the emergence of differences of protein PI 5.2,17.3 KD (e); chaps After dealing with tetraploid P. fortunei leaf protein two-dimensional gel electrophoresis protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b) enhanced after the first weakened, PI 5.2, 16.7KD (f) has been reduced
1外源物质抑制泡桐体外植株再生5-Aza、Chaps、苯丁酸对二倍体和四倍体白花泡桐的叶片芽诱导具有抑制作用,并且随着5-Aza、Chaps、苯丁酸浓度的升高抑制作用增强;5-Aza、Chaps、苯丁酸对二倍体和四倍体白花泡桐的根诱导具有抑制作用,随着5-Aza、Chaps、苯丁酸浓度的升高抑制作用增强。外源物质对二倍体白花泡桐的体外植株再生抑制作用大于四倍体白花泡桐体外植株再生。
2外源物质对幼苗形态变化产生明显影响用5-Aza、Chaps、苯丁酸处理二倍体和四倍体白花泡桐幼苗形态变化存在一定的差异,经5-Aza处理的泡桐出现了芽丛生和植株矮化,苯丁酸处理的泡桐均出现了根部有大量愈伤组织,伴随5-Aza、苯丁酸浓度的增加,幼苗的症状越明显。经Chaps处理的二倍体和四倍体白花泡桐泡桐幼苗形态变化不明显,随着外源物质浓度的增大生长抑制作用增强。
3外源物质对幼苗叶片蛋白质的变化产生了影响经5-Aza、Chaps、苯丁酸处理二倍体和四倍体白花泡桐幼苗进行蛋白质双向电泳发现,等电点、分子量、蛋白质含量在不同外源物质、同种外源物质不同浓度处理的幼苗中存在着差异。经5-Aza处理的二倍体白花泡桐幼苗,蛋白质PI 6.6、22KD(a), PI 6.5、21.8KD(b), PI 5.4、14.5 KD(k)增强表达,蛋白PI 5.2、17.3 KD(e) ,PI 5.2、16.7KD(f)减弱表达;而经5-Aza处理四倍体白花泡桐叶片蛋白质PI 6.6、22KD(a), PI 6.5、21.8KD(b)逐渐渐弱;蛋白质PI 5.2、17.3 KD(e)和PI 5.8、PI 5.2、16.7KD(f)逐渐减弱直至消失。经苯丁酸处理二倍体白花泡桐叶片出现特异蛋白质5.2、17.3 KD(e),5.8、16.5 KD(o);经苯丁酸处理四倍体白花泡桐叶片蛋白质PI 6.6、22KD(a),PI 6.5、21.8KD(b),PI 5.2、16.7KD(f)是差异蛋白质。经Chaps处理白花泡桐叶片蛋白质可以发现,随着Chaps浓度增加蛋白质PI 6.4、20.1 KD(p),PI 6.56、19.8 KD(q)逐渐增强,并且出现了差异蛋白质PI 5.2、17.3 KD(e);经chaps处理四倍体白花泡桐叶片蛋白质双向电泳蛋白PI 6.6、22KD(a), PI 6.5、21.8KD(b)先减弱后增强,PI 5.2、16.7KD(f)一直减小。
The pawlownia is one of our country important fast-growing timber wood tree seeds, is improving our country ecological environment and raises the people’s living standards.,but People are seriously affected the enthusiasm of the cultivation of Paulownia. Paulownia callus induction is as a result of a short time and in the same cycle of callus to split more conducive to study physiology and biochemistry;
To study the transcription of DNA methylation inhibitors (5-Aza) and histone- deacetvlase (4-Phenylbutyric acid, Chaps) on the Growth of Paulownia, the experiment with using two-dimensional gel electrophoresis analysis of proteins after 5-Aza, 4-Phenylbutyric acid, Chaps treatment on P. fortunei diploid and tetraploid P. fortunei differences in gene expression product; study of 5-Aza, 4-Phenylbutyric acid ,Chaps, in cell dedifferentiation process role, and then explore the DNA methylation inhibitor (5-Aza) and histone-deacetvlase acetyltransferase inhibitor (4-Phenylbutyric acid, Chaps) of Paulownia leaf bud induction, seedling morphology, protein expression results, Learn eventually 5-Aza, 4-Phenylbutyric acid, Chaps on the growth and development of Paulownia. The main findings are as follows:
1 Inhibition of exogenous substances in vitro plant regeneration of Paulownia 5-Aza, Chaps, 4-Phenylbutyric acid, treatment P. fortunei diploid and tetraploid and tetraploid has in vitro inhibitory effect of plant regeneration, and with 5 -Aza, Chaps, 4-Phenylbutyric acid, enhanced inhibitory concentration increased.
2 Exogenous substances on seedling have effect on morphological changes With 5-Aza, Chaps, 4-Phenylbutyric acid reatment on P. fortunei diploid and tetraploid and tetraploid morphology differences must exist, with 5-Aza deal with the Paulownia appeared numerous buds and plant dwarfing, 4-Phenylbutyric acid treatment of both the roots of Paulownia have substantial callus, with 5-Aza, Chaps, 4-Phenylbutyric acid concentration increased, seedlings, the more obvious symptoms. With Chaps paulownia seedlings of three treatment patterns did not change significantly.
3 Exogenous substances on diploid and tetraploid P. fortunei seedlings the leaves have effect on proteins Leaves of seelding by 5-Aza, Chaps, 4-Phenylbutyric acid to deal with found in two-dimensional gel electrophoresis protein isoelectric point, molecular weight, protein content, the same species of different concentrations of exogenous substances to deal with the difference between the presence of seedlings in the treatment of seedlings differences exist. After dealing with 5-Aza diploid P. fortunei seedlings, protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b), PI 5.4,14.5 KD (k) to enhance the expression of protein PI 5.2,17.3 KD (e ), PI 5.2, 16.7KD (f) reduced expression; while dealing with 5-Aza tetraploid P. fortunei leaf protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b) a gradual diminuendo; protein PI 5.2, 17.3 KD (e) and PI 5.8, PI 5.2, 16.7KD (f) until the disappearance of warning. Paulownia leaves handling by the 4-Phenylbutyric acid appear diploid-specific protein 5.2,17.3 KD (e), 5.8,16.5 KD (o); by 4-Phenylbutyric acid handling protein tetraploid P. fortunei leaves PI 6.6, 22KD (a), PI 6.5, 21.8KD (b), PI 5.2, 16.7KD (f) is the difference in protein. Chaps by P. fortunei leaf protein processing can be found, with the increase in protein. Chaps concentration PI 6.4,20.1 KD (p), PI 6.56,19.8 KD (q) gradually increased, and the emergence of differences of protein PI 5.2,17.3 KD (e); chaps After dealing with tetraploid P. fortunei leaf protein two-dimensional gel electrophoresis protein PI 6.6, 22KD (a), PI 6.5, 21.8KD (b) enhanced after the first weakened, PI 5.2, 16.7KD (f) has been reduced
引文
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