蝴蝶兰组织培养技术体系研究
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摘要
本试验研究目的在于探索蝴蝶兰离体快繁的有效途径,采用带腋芽的花梗段来诱导丛生芽,继而进行增殖培养,最后诱导生根,炼苗移栽。试验以单因素和正交设计为主,对结果作方差分析及多重比较,确定了蝴蝶兰组织培养的一系列技术支持,研究结果表明:
1.蝴蝶兰离体培养适宜的外植体为已开花的花梗中部腋芽。经过预处理的外植体用0.1%氯化汞处理13min能达到很好的效果。初代培养腋芽萌发适宜的激素配比为6-BA 10mg/L+NAA 0.25mg/L。
2.改良KC培养基是最适合蝴蝶兰增殖生长的培养基,其次为1/2 MS培养基。NAA和6-BA是蝴蝶兰增殖生长最适宜的生长素和细胞分裂素,二者的最佳配比为6-BA 7mg/L+NAA 0.5mg/L。蝴蝶兰在pH 5.4～6的微酸性条件下增殖效果最好。
3.不同的添加物均有促进蝴蝶兰增殖的作用,叶片匀浆的效果最好,以5.5g/L的用量效果最佳。琼脂的用量在5.5g/L时最适合蝴蝶兰的增殖生长。活性炭能有效防止褐化和促进生长,综合效果最佳。除外植体4℃冷藏24h处理外,其余处理措施均可减轻组织褐化,但效果均不及活性炭。
4.继代培养时,为得到整齐一致,生长健壮的试管苗,应考虑到芽苗的大小,1cm长的为最佳试材。蔗糖能显著促进蝴蝶兰增殖生长,且适宜浓度为30g/L。在蝴蝶兰大规模工厂化组培生产中,可以用优质白砂糖代替蔗糖作为增殖生长的碳源。可用自来水代替蒸馏水以降低成本。
5.1/2改良KC培养基是最适合蝴蝶兰生根的培养基。IBA是蝴蝶兰生根适宜的生长素,最佳浓度为1mg/L。有机添加物香蕉泥、椰汁和活性炭均能促进生根,它们的最佳配比为香蕉泥70g/L+椰汁140mL/L+活性炭2g/L。根匀浆有利于根伸长生长,以5g/L和15g/L效果较好。但加入根匀浆后,根条数减少。培养基中加入适量的蔗糖有利于根快速而健壮的生长,最佳用量为10g/L。炼苗天数以7～10d为宜,蝴蝶兰移栽基质以疏松、透气的水苔效果最好。
To explore the efficient way in vitro culture,the cutting segments of flower-stem-knot were utilized to induce clustered shoots to multiply.Experiment was designed primarily by single factor and orthogonal design.Through variance analysis and multiple comparisons, we found out a series of technical parameter combination for the tissue culture of Phalaenopsis,which provided the scientifical and technical support for its factorization production.The main results were as follows:
1.The best type of explants in vitro was the cutting segments of flower-mid-stem-knot which flowered.The pretreated explants immersed in 0.1%HgCl_2 for 13 minutes was sterilize well.The treatment of 6-BA 10 mg/L+NAA 0.25 mg/L which could guarantee the quality of clustered shoots,was the most suitable hormone matching of clustered shoots inducement.
2.Modified KC was the best culture medium in all the medium treatments,and 1/2 MS medium was the second optimum medium.NAA and 6-BA were the suitable auxins and cytokines respectively in vitro culture of Phalaenopsis,and the best multiplication efficiency was obtained in the treatment of NAA 0.5 mg/L+6-BA 7 mg/L.pH 5.4～6 was the best treatment which could promote the explant multiplication of Phalaenopsis.
3.All organic compounds could promote the explant multiplication of Phalaenopsis. Leaf juice treatment(5.5 g/L)was the best one in all the organic compount treatments. The concentration of agar adapted to multiplication was 5.5 g/L.Active carbon was the best one in all the treatments,which could control browning and promote explant growth. Except explants preserved at 4℃for 24 hours,the other treatments could decrease browning,but the effects were not as good as active carbon.
4.It was important to consider the size of shoots,1 cm which could obtain strong plants,was the best one.The cane sugar could improve the multiplication coefficient significantly,and the suitable concentration was 30 g/L.In practical tissue culture production of Phalaenopsis,the sand sugar could replace the cane sugar as the source on the multiplication growth stage and tap warter could replace distilled warter to reduce the culture cost.
5.1/2 modified KC was the best rooting culture medium.IBA was the suitable auxins for rooting culture of Phalaenopsis.All organic compounds could induce rooting of Phalaenopsis,and the best rate and quality of rooting were obtained in the treatment of 70 g/L banana+140 mL/L coconut juice+2 g/L active carbon.Root juice could promote the root of Phalaenopsis grew longer,5 g/L and 15 g/L were suitable concentrations,but it could decrease the number of root of Phalaenopsis.The cane sugar in the medium with suitable concentrations(10 g/L),could significantly promote the quality of rooting.The suitable acclimatization time was 7～10 days.The well-penetrated substance adapted to the plant transplantation in the initial stage and aquatic weed was the most suitable transplantation substance.
1.蝴蝶兰离体培养适宜的外植体为已开花的花梗中部腋芽。经过预处理的外植体用0.1%氯化汞处理13min能达到很好的效果。初代培养腋芽萌发适宜的激素配比为6-BA 10mg/L+NAA 0.25mg/L。
2.改良KC培养基是最适合蝴蝶兰增殖生长的培养基,其次为1/2 MS培养基。NAA和6-BA是蝴蝶兰增殖生长最适宜的生长素和细胞分裂素,二者的最佳配比为6-BA 7mg/L+NAA 0.5mg/L。蝴蝶兰在pH 5.4～6的微酸性条件下增殖效果最好。
3.不同的添加物均有促进蝴蝶兰增殖的作用,叶片匀浆的效果最好,以5.5g/L的用量效果最佳。琼脂的用量在5.5g/L时最适合蝴蝶兰的增殖生长。活性炭能有效防止褐化和促进生长,综合效果最佳。除外植体4℃冷藏24h处理外,其余处理措施均可减轻组织褐化,但效果均不及活性炭。
4.继代培养时,为得到整齐一致,生长健壮的试管苗,应考虑到芽苗的大小,1cm长的为最佳试材。蔗糖能显著促进蝴蝶兰增殖生长,且适宜浓度为30g/L。在蝴蝶兰大规模工厂化组培生产中,可以用优质白砂糖代替蔗糖作为增殖生长的碳源。可用自来水代替蒸馏水以降低成本。
5.1/2改良KC培养基是最适合蝴蝶兰生根的培养基。IBA是蝴蝶兰生根适宜的生长素,最佳浓度为1mg/L。有机添加物香蕉泥、椰汁和活性炭均能促进生根,它们的最佳配比为香蕉泥70g/L+椰汁140mL/L+活性炭2g/L。根匀浆有利于根伸长生长,以5g/L和15g/L效果较好。但加入根匀浆后,根条数减少。培养基中加入适量的蔗糖有利于根快速而健壮的生长,最佳用量为10g/L。炼苗天数以7～10d为宜,蝴蝶兰移栽基质以疏松、透气的水苔效果最好。
To explore the efficient way in vitro culture,the cutting segments of flower-stem-knot were utilized to induce clustered shoots to multiply.Experiment was designed primarily by single factor and orthogonal design.Through variance analysis and multiple comparisons, we found out a series of technical parameter combination for the tissue culture of Phalaenopsis,which provided the scientifical and technical support for its factorization production.The main results were as follows:
1.The best type of explants in vitro was the cutting segments of flower-mid-stem-knot which flowered.The pretreated explants immersed in 0.1%HgCl_2 for 13 minutes was sterilize well.The treatment of 6-BA 10 mg/L+NAA 0.25 mg/L which could guarantee the quality of clustered shoots,was the most suitable hormone matching of clustered shoots inducement.
2.Modified KC was the best culture medium in all the medium treatments,and 1/2 MS medium was the second optimum medium.NAA and 6-BA were the suitable auxins and cytokines respectively in vitro culture of Phalaenopsis,and the best multiplication efficiency was obtained in the treatment of NAA 0.5 mg/L+6-BA 7 mg/L.pH 5.4～6 was the best treatment which could promote the explant multiplication of Phalaenopsis.
3.All organic compounds could promote the explant multiplication of Phalaenopsis. Leaf juice treatment(5.5 g/L)was the best one in all the organic compount treatments. The concentration of agar adapted to multiplication was 5.5 g/L.Active carbon was the best one in all the treatments,which could control browning and promote explant growth. Except explants preserved at 4℃for 24 hours,the other treatments could decrease browning,but the effects were not as good as active carbon.
4.It was important to consider the size of shoots,1 cm which could obtain strong plants,was the best one.The cane sugar could improve the multiplication coefficient significantly,and the suitable concentration was 30 g/L.In practical tissue culture production of Phalaenopsis,the sand sugar could replace the cane sugar as the source on the multiplication growth stage and tap warter could replace distilled warter to reduce the culture cost.
5.1/2 modified KC was the best rooting culture medium.IBA was the suitable auxins for rooting culture of Phalaenopsis.All organic compounds could induce rooting of Phalaenopsis,and the best rate and quality of rooting were obtained in the treatment of 70 g/L banana+140 mL/L coconut juice+2 g/L active carbon.Root juice could promote the root of Phalaenopsis grew longer,5 g/L and 15 g/L were suitable concentrations,but it could decrease the number of root of Phalaenopsis.The cane sugar in the medium with suitable concentrations(10 g/L),could significantly promote the quality of rooting.The suitable acclimatization time was 7～10 days.The well-penetrated substance adapted to the plant transplantation in the initial stage and aquatic weed was the most suitable transplantation substance.
引文
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