苋菜离体培养与试管开花以及幼苗生长的微重力效应
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摘要
本试验以苋菜(Amaranthus mangostanus L)种子为材料,进行离体繁殖及试管开花研究,比较环境因子对苋菜试管苗色素累积以及可溶性蛋白和MDA含量变化的影响,并探讨苋菜幼苗生长和光合色素变化的微重力效应。主要研究结果如下:
1苋菜离体再生体系的建立
以苋菜种子为材料,进行苋菜离体繁殖研究。结果表明:苋菜种子经过75%酒精消毒1min,再用升汞消毒8min,在MS培养基上萌发率可达87%以上。从无菌苗切取的带2个子叶的下胚轴,在MS+6-BA3.0mg/L+NAA0.2mg/L培养基上诱导不定芽效果最好;不定芽在MS+6-BA 3.0 mg/L +IAA1.0mg/L培养基上,增殖效果最好,增殖系数可达13.02。试管苗在1/2MS+0.2 mg/LIBA培养基上生根效果较好,生根率可达20.35%。在适宜温度下,在菜园土上试管苗移栽成活率可达92.5%以上。
2苋菜试管开花
在建立苋菜高效离体再生体系的基础上,研究了苋菜试管开花的影响因子。结果表明:基本培养基MS和1/2MS对苋菜试管开花的影响不大。外源激素对苋菜试管开花的研究表明,单独添加NAA,可促进苋菜试管开花;苋菜试管开花对IBA的浓度变化较敏感;6-BA浓度1.0mg/L时试管苗开花率最高,培养50-60d时可达58.7%。不同单色光处理时,绿光与对照白光试管开花率差异不显著,蓝光、红光处理降低试管苗开花率。每天光照18h可促进苋菜试管开花,培养50-60d时开花率达到59.52%。不同pH处理时,pH5.8试管苗开花率最高。外植体苗龄在15-21d时试管苗开花率差异不显著,较适宜诱导成花,苗龄超过这个范围时试管开花率都有所下降;继代可抑制苋菜试管开花。不继代连续培养3个月以上苋菜试管苗开花率可以达到95%以上。
离体诱导的花发育正常,可以在试管内授粉受精,并获得有萌发力的种子,有时甚至可以在试管内直接萌发成苗。
苋菜试管苗由营养生长向生殖生长的过程中,蛋白质含量、可溶性糖含量以及苋菜红色素含量出现一个升高和突降的过程,结合观察结果推测,苋菜花芽分化过程可能发生在初次转接后培养19-28d这一阶段。
3环境因子对苋菜试管苗色素累积的影响
温度、pH、光质和光周期等条件对苋菜光合色素、苋菜红色素以及花色素苷含量变化的影响研究表明:温度20±1℃,pH5.4,白光照射以及弱光条件下,每天光照24 h均可促进叶绿素的合成;类胡萝卜素对环境条件的响应表现与叶绿素一致;温度范围20-32℃时对苋菜花色素苷含量变化差异不显著,pH5.4和pH5.8时花色苷含量均较高,白光对花色素苷累积效果较好,光照18h/d和12h/d花色素苷相对含量较高且差异不显著;20±1℃时苋菜红色素含量较高。
4环境因子对苋菜试管苗可溶性蛋白和MDA变化的影响
温度、pH、光质和光周期等条件对苋菜试管苗可溶性蛋白和丙二醛含量变化的影响研究表明:较低温度使苋菜试管苗可溶性蛋白含量升高,20±1℃时可溶性蛋白含量最高,pH6.2和pH6.4时可溶性蛋白含量均高于其他处理,且差异不显著,红光可促进可溶性蛋白合成,弱光24h/d时可溶性蛋白含量最高;蓝光照射时MDA含量最低,光周期6h/d和12h/d时MDA含量差异不显著,高于其他处理,pH6.7时丙二醛含量最高。
5苋菜幼苗生长的微重力效应
以湿润培养为对照研究了模拟微重力效应培养对苋菜幼苗生长以及光和色素含量的影响。结果表明:微重力效应可以使苋菜幼苗的高度增加,模拟微重力培养40d后苋菜幼苗的叶绿素含量下降14.12%,类胡萝卜素下降11.68%。
Seeds were used to study in vitro propagation and in vitro flowering of amaranth. The effects of environmental factors on the contents of pigment, soluble proteins and MDA were compared. And the effects of microgravity on the seedling growth and the contents of photosynthetic pigment were discussed. The main results were as follows:
1 Establishment of in vitro regeneration system of amaranth
Seeds were used to study in vitro propagation of amaranth. The results showed that the germination rate reached 87% or more on the MS medium after the sterilization treatment of 75% alcohol for 1min and 0.1% mercuric chloride for 8min. The best medium for adventitious bud induction from the hypocotyl with two cotyledons was MS medium with addition of 3.0 mg/L 6-BA, 0.2mg/L NAA, for propagation was MS medium with addition of 3.0mg/L 6-BA, 1.0mg/L IAA, with the proliferation coefficient of 13.02, for rooting was 1/2MS medium with addition of 0.2mg/L IBA, with the rooting rate of 20.35%. When the plantlets were transferred onto the garden soil, and the survival rate was up to 92.5% at proper temperature.
2 In vitro flowering of amaranth
The affecting factors of in vitro flowering were studied on the establishment of efficient regeneration system. Based on the results, the in vitro flowering rate of amaranth had little difference between MS and 1/2MS basal media. Phytohormones had effects on the development of flowering. When either NAA or IBA alone was used, it could promote the in vitro flowering. When the concentration of 6-BA was 1.0mg / L, the in vitro flowering was the best, and the flowering rate was up to 58.7% after 50-60 days. In vitro flowering rate was no significant difference between the green light and white light treatments, but reduced by blue and red light treatments. The in vitro flowering was promoted under the light of 18h/d, and the flowering rate was up to 59.52% after 50-60d. The flowering rate was highest under pH5.8. The in vitro flowering rate was no significant difference when the seedling age of the explants at 15-21days; otherwise it decreased if seedling age was over this range. Without the next generations and continuously cultured for more than 3 months, the flowering rate could reach more than 95%.
The flowers induced from the in vitro plantlets developed well and the fertilization and seed development were normal, and sometime the seeds even germinated into seedlings in the culturing bottle.
The content of soluble protein, soluble sugar and the amaranth red pigment presented the tendency of increasing firstly, then decreasing during the plantlets transformed from vegetative growth to reproductive growth. The flower bud differentiation of amaranth maybe happened after the first next generations 19-28d.
3 The effects of environmental factors on the accumulation of pigment of in vitro amaranth plantlets
The effects of the different environmental factors, such as temperature, pH value, light quality and photoperiod, were studied on the accumulation of pigment of in vitro amaranth plantlets. The results showed that both the chlorophyll synthesis and carotenoid synthesis were increased at 20±1℃, with pH 5.4, white light under 24 h/d. The content of anthocyanin was no significant difference at 20-32℃, higher between pH5.4 and pH5.8, higher through white light treatments, higher and not significant difference between 12 h/d and 18 h/d. The content of amaranth red pigment was higher at 20±1℃.
4 The effects of environmental factors on the contents of soluble proteins and MDA of amaranth plantlets
The effects of the different environmental factors, such as temperature, pH values, light quality and photoperiod, were studied on the contents of soluble proteins and MDA of amaranth Test-tube seedlings. The research results showed that the contents of soluble proteins of amaranth plantlets increased at lower temperature and highest at 20±1℃. The content of soluble proteins was higher and not significant difference between pH6.2 and pH6.4. The soluble protein synthesis was promoted through red light treatment. The content of soluble proteins was highest with low-light under 24 h/d. The content of MDA was lowest through blue light treatment, was higher and not significant difference between 6h h/d and 12 h/d, and was highest at pH6.7.
5 The simulate microgravity effects on amaranth seedling growth
The effects of simulate microgravity on the growth of amaranth seedlings and the content of photosynthetic pigment were studied. The results showed that the height of amaranth seedlings increased and the content of photosynthetic pigment decreased through the simulate microgravity effects. The content of chlorophyll decreased by 14.12% and carotenoid decreased by 11.68% after 40d simulated microgravity treatment.
1苋菜离体再生体系的建立
以苋菜种子为材料,进行苋菜离体繁殖研究。结果表明:苋菜种子经过75%酒精消毒1min,再用升汞消毒8min,在MS培养基上萌发率可达87%以上。从无菌苗切取的带2个子叶的下胚轴,在MS+6-BA3.0mg/L+NAA0.2mg/L培养基上诱导不定芽效果最好;不定芽在MS+6-BA 3.0 mg/L +IAA1.0mg/L培养基上,增殖效果最好,增殖系数可达13.02。试管苗在1/2MS+0.2 mg/LIBA培养基上生根效果较好,生根率可达20.35%。在适宜温度下,在菜园土上试管苗移栽成活率可达92.5%以上。
2苋菜试管开花
在建立苋菜高效离体再生体系的基础上,研究了苋菜试管开花的影响因子。结果表明:基本培养基MS和1/2MS对苋菜试管开花的影响不大。外源激素对苋菜试管开花的研究表明,单独添加NAA,可促进苋菜试管开花;苋菜试管开花对IBA的浓度变化较敏感;6-BA浓度1.0mg/L时试管苗开花率最高,培养50-60d时可达58.7%。不同单色光处理时,绿光与对照白光试管开花率差异不显著,蓝光、红光处理降低试管苗开花率。每天光照18h可促进苋菜试管开花,培养50-60d时开花率达到59.52%。不同pH处理时,pH5.8试管苗开花率最高。外植体苗龄在15-21d时试管苗开花率差异不显著,较适宜诱导成花,苗龄超过这个范围时试管开花率都有所下降;继代可抑制苋菜试管开花。不继代连续培养3个月以上苋菜试管苗开花率可以达到95%以上。
离体诱导的花发育正常,可以在试管内授粉受精,并获得有萌发力的种子,有时甚至可以在试管内直接萌发成苗。
苋菜试管苗由营养生长向生殖生长的过程中,蛋白质含量、可溶性糖含量以及苋菜红色素含量出现一个升高和突降的过程,结合观察结果推测,苋菜花芽分化过程可能发生在初次转接后培养19-28d这一阶段。
3环境因子对苋菜试管苗色素累积的影响
温度、pH、光质和光周期等条件对苋菜光合色素、苋菜红色素以及花色素苷含量变化的影响研究表明:温度20±1℃,pH5.4,白光照射以及弱光条件下,每天光照24 h均可促进叶绿素的合成;类胡萝卜素对环境条件的响应表现与叶绿素一致;温度范围20-32℃时对苋菜花色素苷含量变化差异不显著,pH5.4和pH5.8时花色苷含量均较高,白光对花色素苷累积效果较好,光照18h/d和12h/d花色素苷相对含量较高且差异不显著;20±1℃时苋菜红色素含量较高。
4环境因子对苋菜试管苗可溶性蛋白和MDA变化的影响
温度、pH、光质和光周期等条件对苋菜试管苗可溶性蛋白和丙二醛含量变化的影响研究表明:较低温度使苋菜试管苗可溶性蛋白含量升高,20±1℃时可溶性蛋白含量最高,pH6.2和pH6.4时可溶性蛋白含量均高于其他处理,且差异不显著,红光可促进可溶性蛋白合成,弱光24h/d时可溶性蛋白含量最高;蓝光照射时MDA含量最低,光周期6h/d和12h/d时MDA含量差异不显著,高于其他处理,pH6.7时丙二醛含量最高。
5苋菜幼苗生长的微重力效应
以湿润培养为对照研究了模拟微重力效应培养对苋菜幼苗生长以及光和色素含量的影响。结果表明:微重力效应可以使苋菜幼苗的高度增加,模拟微重力培养40d后苋菜幼苗的叶绿素含量下降14.12%,类胡萝卜素下降11.68%。
Seeds were used to study in vitro propagation and in vitro flowering of amaranth. The effects of environmental factors on the contents of pigment, soluble proteins and MDA were compared. And the effects of microgravity on the seedling growth and the contents of photosynthetic pigment were discussed. The main results were as follows:
1 Establishment of in vitro regeneration system of amaranth
Seeds were used to study in vitro propagation of amaranth. The results showed that the germination rate reached 87% or more on the MS medium after the sterilization treatment of 75% alcohol for 1min and 0.1% mercuric chloride for 8min. The best medium for adventitious bud induction from the hypocotyl with two cotyledons was MS medium with addition of 3.0 mg/L 6-BA, 0.2mg/L NAA, for propagation was MS medium with addition of 3.0mg/L 6-BA, 1.0mg/L IAA, with the proliferation coefficient of 13.02, for rooting was 1/2MS medium with addition of 0.2mg/L IBA, with the rooting rate of 20.35%. When the plantlets were transferred onto the garden soil, and the survival rate was up to 92.5% at proper temperature.
2 In vitro flowering of amaranth
The affecting factors of in vitro flowering were studied on the establishment of efficient regeneration system. Based on the results, the in vitro flowering rate of amaranth had little difference between MS and 1/2MS basal media. Phytohormones had effects on the development of flowering. When either NAA or IBA alone was used, it could promote the in vitro flowering. When the concentration of 6-BA was 1.0mg / L, the in vitro flowering was the best, and the flowering rate was up to 58.7% after 50-60 days. In vitro flowering rate was no significant difference between the green light and white light treatments, but reduced by blue and red light treatments. The in vitro flowering was promoted under the light of 18h/d, and the flowering rate was up to 59.52% after 50-60d. The flowering rate was highest under pH5.8. The in vitro flowering rate was no significant difference when the seedling age of the explants at 15-21days; otherwise it decreased if seedling age was over this range. Without the next generations and continuously cultured for more than 3 months, the flowering rate could reach more than 95%.
The flowers induced from the in vitro plantlets developed well and the fertilization and seed development were normal, and sometime the seeds even germinated into seedlings in the culturing bottle.
The content of soluble protein, soluble sugar and the amaranth red pigment presented the tendency of increasing firstly, then decreasing during the plantlets transformed from vegetative growth to reproductive growth. The flower bud differentiation of amaranth maybe happened after the first next generations 19-28d.
3 The effects of environmental factors on the accumulation of pigment of in vitro amaranth plantlets
The effects of the different environmental factors, such as temperature, pH value, light quality and photoperiod, were studied on the accumulation of pigment of in vitro amaranth plantlets. The results showed that both the chlorophyll synthesis and carotenoid synthesis were increased at 20±1℃, with pH 5.4, white light under 24 h/d. The content of anthocyanin was no significant difference at 20-32℃, higher between pH5.4 and pH5.8, higher through white light treatments, higher and not significant difference between 12 h/d and 18 h/d. The content of amaranth red pigment was higher at 20±1℃.
4 The effects of environmental factors on the contents of soluble proteins and MDA of amaranth plantlets
The effects of the different environmental factors, such as temperature, pH values, light quality and photoperiod, were studied on the contents of soluble proteins and MDA of amaranth Test-tube seedlings. The research results showed that the contents of soluble proteins of amaranth plantlets increased at lower temperature and highest at 20±1℃. The content of soluble proteins was higher and not significant difference between pH6.2 and pH6.4. The soluble protein synthesis was promoted through red light treatment. The content of soluble proteins was highest with low-light under 24 h/d. The content of MDA was lowest through blue light treatment, was higher and not significant difference between 6h h/d and 12 h/d, and was highest at pH6.7.
5 The simulate microgravity effects on amaranth seedling growth
The effects of simulate microgravity on the growth of amaranth seedlings and the content of photosynthetic pigment were studied. The results showed that the height of amaranth seedlings increased and the content of photosynthetic pigment decreased through the simulate microgravity effects. The content of chlorophyll decreased by 14.12% and carotenoid decreased by 11.68% after 40d simulated microgravity treatment.
引文
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