菊薯愈伤组织诱导及再分化初步研究
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摘要
选择菊薯优良株系为试验材料,以嫩叶、腋芽、冠芽等为外植体,分别对消毒程序、筛选方法、愈伤组织诱导、不定芽分化诱导、继代增殖培养、生根诱导培养等离体快繁过程中的各个环节进行了初步研究。
外植体消毒试验结果表明:不同外植体对0.1%HgCl_2溶液的敏感性存在着一定的差异,但均达到较好的消毒效果。其消毒程序为:流水冲洗2～3h后,用浓度70%～75%酒精表面消毒10～15s后,无菌水冲洗2～3次,再用0.1%HgCl_2消毒8min,无菌水冲洗3～5次。
在外植体筛选和愈伤组织诱导试验中发现,适合无菌苗培养的外植体材料有叶片、腋芽、茎节组织和块茎冠芽等。
研究发现菊薯离体培养分化成苗至少有三种途径:即无菌短枝型、器官发生型、器官直接转变型。
研究结果表明,适合菊薯外植体材料诱导愈伤组织的最佳培养基是:MS+6-BA1～1.5 mg·L~(-1)+NAA0.5～1.0 mg·L~(-1)+3%蔗糖+0.7%琼脂,pH值5.8。最适宜的诱导培养时间为:7～10d。。
不定芽分化诱导的研究结果表明,来源不同的愈伤组织对不同激素和配比所表现出的分化诱导效应不同,进行不定芽诱导的最佳培养基是:MS+6-BA1.0 mg·L~(-1)+NAA 0.1 mg·L~(-1)+3%蔗糖+0.7%琼脂,pH值5.8,诱导分化率达79.28%.最适宜的诱导培养时间为60～80d。
在继代增殖培养研究中发现:菊薯愈伤组织在继代培养时极易发生褐化而死亡;6-BA、NAA、GA_3对菊薯不定芽继代增殖影响的主次顺序是6-BA、GA_3和NAA;对不定芽生长影响的主次顺序是GA_3、6-BA、NAA;不定芽继代增殖效果最佳的培养基为:MS+6-BA1.0mg·L~(-1)+NAA0.01 mg·L~(-1)+GA_30.1mg·L~(-1),增殖率为567%。
生根诱导试验结果表明:菊薯试管苗具有极易生根的特性。大量元素对生根率具有一定的影响,以1/2MS+NAA0.01 mg·L~(-1)作为生根培养基最适宜,生根率达到88.89%。不同浓度的为NAA对生根诱导率影响不明显,但具有壮苗和加快生根速度的作用。
The strains of Smallanthus sonchifolius were selected as the experimental materials,Tender leaves,axillary buds and crown buds were used as explants.The explants were studied with surface-sterillized with 0.1%HgCl_2,method for screening,callus induction, induction of adventitious bud differentiation,subculture and rooting induction of plantlets,etc. The conclusions were as follows:
Study on disinfecting methods of explant.The experimental results show that the sensitivity of explant on 0.1%HgCl_2 was not the same,but the disinfection effects were good. The disinfection processes as follows:The explants were washed in water for 2～3hours, before putting in 70%～75%ethanol for 10～15s and rinsed 3～5 times with sterile deionized water,then the explants were surface sterilized with 0.1%HgCl_2 for 8mins and rinsed 3～5 times with sterile deionized water.
Study on induction tests of explant and callus.The results showed that the most suitable explant materials are leaves,axillary buds,tubers crown buds,etc.
The study found that there are at least three ways of seedlings differenttiation of explants of Smallanthus sonchifolius,namely,aseptic cuttage type,organogenesis type,organ type.
The study results of inducing callus of explant showed that The optimum medium for inducing callus is MS with 1.0～1.5 mg·L~(-1)6-BA+0.5～1.0 mg·L~(-1)IAA,3%sucrose and 0.6% agar,pH5.8.the most appropriate time of inducing callus is 7～10 days
The study results of inducing adventitious bud showed that induced effects callus of different explants on different hormones and the ratio are not the same as.The optimum medium of inducing adventitious buds is MS with 1.0 mg·L~(-1)6-BA +0.1 mg·L~(-1)IAA,3% sucrose and 0.6%agar,pH5.8,and the inducing rate is 79.28%.the most appropriate time for inducing adventitious buds is 60～80 days
The results for multiplication of adventitious buds show that the callus browning and death can easily happen at the proliferation of cultured.6-BA,NAA,GA3 to the proliferation of adventitious buds the impact of the order are 6-BA,GA3 and NAA.6-BA,NAA,GA3 to the growth the impact of the order are 6-BA,GA3 and NAA.The optimum medium for proliferation of adventitious buds is MS with 1.0 mg·L~(-1)6-BA+0.1 mg·L~(-1)IAA,3%sucrose and 0.6%agar,pH5.8,and the proliferation rate is 567%.
The experimental results for rooting induction show that plantlet of Smallanthus sonchifolius is easily rooting,Major element in medium was more effective in inducing rooting,1/2MS+NAA0.01 mg·L~(-1)was optimum for rooting,with rooting percentage more than 88.89%.NO obvious effect on the rooting of the different concentrations of NAA was seen, but The NAA could certainly promote the strong seedling,and can sped up the rooting speed.
外植体消毒试验结果表明:不同外植体对0.1%HgCl_2溶液的敏感性存在着一定的差异,但均达到较好的消毒效果。其消毒程序为:流水冲洗2～3h后,用浓度70%～75%酒精表面消毒10～15s后,无菌水冲洗2～3次,再用0.1%HgCl_2消毒8min,无菌水冲洗3～5次。
在外植体筛选和愈伤组织诱导试验中发现,适合无菌苗培养的外植体材料有叶片、腋芽、茎节组织和块茎冠芽等。
研究发现菊薯离体培养分化成苗至少有三种途径:即无菌短枝型、器官发生型、器官直接转变型。
研究结果表明,适合菊薯外植体材料诱导愈伤组织的最佳培养基是:MS+6-BA1～1.5 mg·L~(-1)+NAA0.5～1.0 mg·L~(-1)+3%蔗糖+0.7%琼脂,pH值5.8。最适宜的诱导培养时间为:7～10d。。
不定芽分化诱导的研究结果表明,来源不同的愈伤组织对不同激素和配比所表现出的分化诱导效应不同,进行不定芽诱导的最佳培养基是:MS+6-BA1.0 mg·L~(-1)+NAA 0.1 mg·L~(-1)+3%蔗糖+0.7%琼脂,pH值5.8,诱导分化率达79.28%.最适宜的诱导培养时间为60～80d。
在继代增殖培养研究中发现:菊薯愈伤组织在继代培养时极易发生褐化而死亡;6-BA、NAA、GA_3对菊薯不定芽继代增殖影响的主次顺序是6-BA、GA_3和NAA;对不定芽生长影响的主次顺序是GA_3、6-BA、NAA;不定芽继代增殖效果最佳的培养基为:MS+6-BA1.0mg·L~(-1)+NAA0.01 mg·L~(-1)+GA_30.1mg·L~(-1),增殖率为567%。
生根诱导试验结果表明:菊薯试管苗具有极易生根的特性。大量元素对生根率具有一定的影响,以1/2MS+NAA0.01 mg·L~(-1)作为生根培养基最适宜,生根率达到88.89%。不同浓度的为NAA对生根诱导率影响不明显,但具有壮苗和加快生根速度的作用。
The strains of Smallanthus sonchifolius were selected as the experimental materials,Tender leaves,axillary buds and crown buds were used as explants.The explants were studied with surface-sterillized with 0.1%HgCl_2,method for screening,callus induction, induction of adventitious bud differentiation,subculture and rooting induction of plantlets,etc. The conclusions were as follows:
Study on disinfecting methods of explant.The experimental results show that the sensitivity of explant on 0.1%HgCl_2 was not the same,but the disinfection effects were good. The disinfection processes as follows:The explants were washed in water for 2～3hours, before putting in 70%～75%ethanol for 10～15s and rinsed 3～5 times with sterile deionized water,then the explants were surface sterilized with 0.1%HgCl_2 for 8mins and rinsed 3～5 times with sterile deionized water.
Study on induction tests of explant and callus.The results showed that the most suitable explant materials are leaves,axillary buds,tubers crown buds,etc.
The study found that there are at least three ways of seedlings differenttiation of explants of Smallanthus sonchifolius,namely,aseptic cuttage type,organogenesis type,organ type.
The study results of inducing callus of explant showed that The optimum medium for inducing callus is MS with 1.0～1.5 mg·L~(-1)6-BA+0.5～1.0 mg·L~(-1)IAA,3%sucrose and 0.6% agar,pH5.8.the most appropriate time of inducing callus is 7～10 days
The study results of inducing adventitious bud showed that induced effects callus of different explants on different hormones and the ratio are not the same as.The optimum medium of inducing adventitious buds is MS with 1.0 mg·L~(-1)6-BA +0.1 mg·L~(-1)IAA,3% sucrose and 0.6%agar,pH5.8,and the inducing rate is 79.28%.the most appropriate time for inducing adventitious buds is 60～80 days
The results for multiplication of adventitious buds show that the callus browning and death can easily happen at the proliferation of cultured.6-BA,NAA,GA3 to the proliferation of adventitious buds the impact of the order are 6-BA,GA3 and NAA.6-BA,NAA,GA3 to the growth the impact of the order are 6-BA,GA3 and NAA.The optimum medium for proliferation of adventitious buds is MS with 1.0 mg·L~(-1)6-BA+0.1 mg·L~(-1)IAA,3%sucrose and 0.6%agar,pH5.8,and the proliferation rate is 567%.
The experimental results for rooting induction show that plantlet of Smallanthus sonchifolius is easily rooting,Major element in medium was more effective in inducing rooting,1/2MS+NAA0.01 mg·L~(-1)was optimum for rooting,with rooting percentage more than 88.89%.NO obvious effect on the rooting of the different concentrations of NAA was seen, but The NAA could certainly promote the strong seedling,and can sped up the rooting speed.
引文
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