Ethanol Metabolism in Calluses of Several Selected Plant Species on Two Typical Plant-Growth-Regulator Balanced Media

For investigation on the characteristics of ethanol metabolism in tissues of different plant species, calluses from eight selected plant species were cultured on medium supplemented with ethanol in tightly sealed culture flasks. Changes of the ethanol level were detected by gas chromatography. During the culture period, the calluses of tobacco, potato and petunia were, able to catabolize exogenous ethanol, resulting in the prominent decline of the ethanol level in the medium. The calluses of melon and peanut were also able to catabolize thanol but with lower efficiency. The other three calluses of carrot, soybean and rice did not catabolize ethanol but instead produced small to large amount of ethanol, resulting in the increase of the ethanol level in the media. It was also found that changing the balance between auxin and cytokinin could influence only the ethanol metabolism efficiency but could not change the metabolism patterns on ethanol of the cultured calluses. It can be concluded that, ethanol metabolism pattern of calluses in cultures is an innate physiological characteristic of the respective plant species.

Influences of Carbon Sources and Plant Growth Regulators on Anther Culture Efficiency of Pepper

[Objective]In order to increase anther culture efficiency of pepper.[Method]MS culture media and Bolajiaohong were used in this experiment to study the influences of carbon sources and concentrations on anther callus induction of pepper.Jiayu was taken as a material to study influences of plant growth regulators and concentrations on anther callus induction of pepper according to L16（4^5） orthogonal design.[Result]The average callus and embryoid induction rates of maltose at all concentrations were higher than these of sucrose but the difference was not significant.Taking maltose or sucrose as a carbon source,3% to 6% concentration was good for increasing induction frequencies of calli and embryoids.However,If the concentration was over 6%,the induction rates were declined dramatically with the increase of sugar concentration.The influences of growth regulators on induction rate of calli were listed as 2,4-D﹥ZT﹥NAA﹥KT﹥6-BA;the influences on induction rates of embryoids were listed as 2,4-D﹥NAA﹥ZT﹥KT﹥6-BA.The 2,4-D,ZT,NAA and KT had signficant or extremely significant influences on induction rates of calli and embryoids.2,4-D,ZT at 1.0 mg/L and NNA,KT at 0.5 mg/L had the best effects.The influences of ZT on calli and embryoids were better than those of KT and 6-BA.1.0 mg/L 2,4-D ＋1.0 mg/L ZT ＋0.5 mg/L KT ＋0.5 mg/L 6-BA was the best regulator combination for induction culture of Jiayu anther.[Conclusion]The experiment provided research basis for anther culture of pepper.

Influencing in vitro clonal propagation of Chonemorpha fragrans(moon) Alston by culture media strength,plant growth regulators,carbon source and photo periodic incubation

Chonemorpha fragrans is an endangered medicinal woody climber,regarded among alternative plant sources of camptothecin.Camptothecin is a monoterpene indole anti-cancer alkaloid with annual trade value of over three billion U.S.dollars in the recent,and is used in the production of its analog drugs approved for the chemotherapy of cancer of varied types.Effects of plant growth regulators,culture media strength and photoperi-odic duration on the micropropagation ef ciency of C.fragrans from nodal segment explants were studied on Murashige and Skoog(MS)medium amended with Thidiazuron(TDZ),Benzylaminopurine(BAP)or Kinetin(Kin).Thidiazuron was more ef cient over BAP and Kin when half basal MS medium was used over full or quarter strength.Results of carbon source experiment showed sucrose as the most effective over glucose,fructose,and maltose in the clonal production.Studies on the photope-riodic incubation duration showed 12 h as the best light period and sub or supra-optimal resulted in the production of abnormal and albino micro shoots.Experimental results on the evaluation of physiological,biochemical parameters showed the role of pigment molecules and antioxidant systems in the production of albino micro shoots.

Effects of variations in culture media and hormonal treatments upon callus induction potential in endosperm explant of Barringtonia racemosa L.

Objective: To induce callus from the medicinally valuable species, Barringtonia racemosa L.(B. racemosa) whereby the formation of callus is essential for micropropagation studies and in vitro plant secondary metabolites production.Methods: The callus induction potential in B. racemosa was assessed from endosperm explant cultured on different culture media and plant hormonal treatments. Lloyd and Mc Cown's woody plant medium and Murashige and Skoog's medium were used in the study as culture media. On the other hand, various concentrations and combinations of2,4-dichlorophenoxyacetic acid(1.0–2.0 mg/L) and kinetin(0.5–2.5 mg/L) had been incorporated in the culture media to exert the effects of auxin and cytokinin on callus induction.Results: From the present study, it was found that the profuse [(1.681 ± 0.770) g fresh weight,(0.239 ± 0.239) g dry weight] and friable callus formation was optimally produced with desirable morphology and considerable percentage of callus induction(56.70%) in endosperm explants cultured on 1.0 mg/L 2,4-dichlorophenoxyacetic acid and 1.5 mg/L kinetin in Murashige and Skoog's medium.Conclusions: A reliable protocol for inducing callus formation of profuse and friable morphology in endosperm explant of B. racemosa had therefore been successfully established.

High Frequency Plant Regeneration from Leaf Derived Callus of <i>Dianthus caryophyllus</i>L.

An efficient procedure was developed for in vitro callus induction, proliferation and regeneration of carnation cultivar (Dianthus caryophyllus L.) using leaf, nodal and inter-nodal explants on Murashige and Skoog’s medium (MS) supplemented with exogenous plant growth regulators. For morphogenic callus induction and proliferation from various explants, MS medium supplemented with 3.0 mg/l 2,4-D was highly efficient with 100% callus induction frequency from inter-nodal explants. Leaf explants showed quicker response than nodal and inter-modal explants, for callus initiation within 6 days of inoculation. Best grown callus was obtained from leaf explant. The leaf-derived callus was maintained up to several weeks, which indicated that 8-week incubation period was the most suitable for obtaining well proliferated, morphogenic callus. Temperature variation also affected the growth of in vitro induced morphogenic callus from various explants. Results have shown that 27°C proved to be the best temperature for morphogenic callus induction and proliferation from leaf and inter-nodal explants. Among the auxin-cytokinin combination, MS medium containing 1.0 mg/l N(6)-benzylaminopurin (BAP) and 2.0 mg/l NAA showed the highest efficiency of callus initiation and proliferation from leaf, nodal and inter-nodal explants. Light conditions proved better for callogenesis and proliferation from leaf, nodal and inter-nodal explants. Regeneration response from well grown morphogenic callus was prominent on MS medium supplemented with 3.0 mg/l BAP alone and 1.0 mg/l NAA with 3.0 mg/l BAP.

Plant Regeneration Through Indirect Somatic Embryogenesis in Coelogyne Cristata Orchid

A newly efficient protocol has been established for high frequency somatic embryogenesis through callus culture of Coelogyne cristata. The best frequency of callusing was obtained from leaf segments (3-5 mm) cultured on the MS medium supplemented with 2 mg?L–1 2,4-D and 2 mg?L–1 BA combination. A negative effect of coconut water was ob-served on the callus induction medium. When callus lines no. 4, 6 and 8 induced from leaf segments were sub-cultured separately on ? MS and MS media containing AC (1-3 g?L–1), formation of somatic embryos was found. However, percentages of embryo formation and the number of embryos per explants were strongly affected by media and callus lines used. The effect of 1/2 MS media is definitely better than MS medium for somatic embryogenesis from the selected lines of leaf derived callus. Among the callus lines, line no. 4 is the best for somatic embryogenesis followed by line no. 6 and 8. The somatic embryos converted into healthy plants with well developed shoots on the same media. The plant-lets were transferred to 1/2 MS medium containing 1 g?L–1 AC for plant regeneration until 8 weeks of culture and successfully acclimatized in the greenhouse.

<i>In vitro</i>propagation of <i>Aegle marmelos</i>(L.) corr., a medicinal plant through axillary bud multiplication

A protocol for rapid in vitro propagation from nodal explants of the medicinal tree species Aegle marmelos (L.) corr. of family Rutaceae has been described. High frequency bud break were induced on Murashige and Skoog’s (1962) basal medium supplemented with 0.5 mg benzyladenine (BA)/l. After 10 days of culture, nodal explants with multiplied buds started callusing with restricted growth and defoliation. When the same nodal explants ware transferred into the same basal medium supplemented with 0.5 mg BA/l with different concentrations of either kinetin (KN) or gibberellic acid (GA3) or in combinations has shown healthy shoots with expanded shoot length. Excised shoots (2cm-3cm long with 2 to 3 nodes) when grown on 1/2 MS basal medium with 2.5 mg/l indole-3-butyric acid (IBA) and 0.5% activated charcoal (A.C.)/l has shown rhizogenesis. After excision, in the second passage, the nodal explants also showed bud break when sub cultured on MS basal medium supplemented with 0.5 mg BA/l. These shoots also successfully rooted on the same above said medium.

Plant Regeneration through Indirect Organogenesis in Two Cultivars of Pineapple (Ananas comosus L.)

Unavailability of performant planting material of pineapple constitutes a major problem of its cultivation in Africa. For this purpose, indirect organogenesis technique is used to evaluate the in vitro responses of two cultivars of pineapple during the explant’s regeneration. Calli were induced from crown leaf and plantlets leaf of “Smooth Cayenne” and “Sugarloaf cultivars”. Murashige and Skoog medium with vitamins B5 supplemented with different growth regulators combinations were used. BAP and/or 2,4-D have been added to base medium for calli cells’ differentiation while BAP and GA3 have been added for plant elongation. The results indicated that explants from regenerated plantlets leaves cultivated on MS supplemented with copper (II) sulphate 5-hydrate concentrations incorporated had significant (p < 0.0001) influence on callus induction in pineapple cultivars. Likewise, MS medium with NAA (0.5 mg/l) + BAP (1 mg/l) had a highly significant influence with 8.8 differentiated Calli. Also, MS medium supplemented with BAP (3 mg/l) + GA3 (2 mg/l) for the “smooth Cayenne” had significantly influenced (p < 0.0001) Calli regeneration with a high rate of 55.25% plantlets. MS medium containing 0.5 mg/l of NAA + 0 mg/l IBA produced a high number of roots in Sugarloaf whereas the medium containing 1.5 mg/l NAA + 0.5 mg/l (IBA) produced high number of roots in smooth Cayenne. We have established an efficient and reproducible protocol for mass propagation and genetic transformation of pineapple though indirect organogenesis. This protocol may be used in genetics engineering studies for pineapple breeding.

生长素和细胞分裂素诱导植物胚性愈伤组织研究进展

植物胚性愈伤组织在植物研究与应用中是一种重要的试材,有很多关于植物胚性愈伤组织诱导方法研究的报道。生长素和细胞分裂素被广泛应用于胚性愈伤组织的诱导,是胚性愈伤组织诱导的关键因子。阐述了生长素和细胞分裂素在植物胚性愈伤组织诱导及体细胞胚胎发生过程中的作用,讨论了其在植物胚性愈伤组织诱导中的作用、使用方法与原则,为愈伤组织诱导、愈伤组织胚性化诱导等研究中植物生长调节剂的使用提供依据。

油莎豆愈伤组织诱导及再生体系的建立

为建立高效的油莎豆愈伤组织诱导及再生体系,对适用于愈伤组织诱导的外植体(幼根、幼叶、幼叶鞘和分蘖节)进行筛选,并探讨不同植物生长调节剂组合对愈伤组织诱导、增殖和分化的影响。结果表明,油莎豆愈伤组织诱导的最佳外植体为分蘖节;最优愈伤组织诱导的植物生长调节剂组合为2.5 mg/L 2,4-D+0.6 mg/L 6-BA+0.6 mg/L NAA+0.9 mg/L ZT,诱导率可达65.58%;最优愈伤组织增殖的植物生长调节剂组合为0.5 mg/L 2,4-D+0.9 mg/L 6-BA+1.2 mg/L NAA+1.2 mg/L ZT,增殖率可达68.12%;最优愈伤组织分化的植物生长调节剂组合为0.9 mg/L 6-BA+0.6 mg/L NAA,分化率可达66.67%。试验筛选出了适合油莎豆愈伤组织诱导的最佳外植体,并优化了油莎豆愈伤组织诱导、增殖和分化的条件,为后续油莎豆体细胞杂交、遗传转化等生物技术的研究及育种应用奠定基础。

不同激素配比对矩镰荚苜蓿4种外植体愈伤组织诱导的影响

【目的】筛选出矩镰荚苜蓿(Medicago archiducis-nicolai)最佳外植体及最佳再生条件。【方法】以矩镰荚苜蓿种子为材料,选择MS(含0.8%琼脂+3%蔗糖)为基本培养基,添加不同种类及浓度配比生长调节剂对矩镰荚苜蓿子叶、叶柄、茎、叶片进行诱导愈伤和愈伤再分化。【结果】外植体诱导率由高到低依次为:茎>叶柄>真叶>子叶,外植体褐化率由高到低依次为:茎<真叶<叶柄<子叶;在1.5 mg/L 2,4-D+0.1 mg/L KT+MS+0.8%琼脂+3%蔗糖条件下最适合真叶诱导愈伤,诱导率为78%,褐化率为8%;1.5 mg/L 2,4-D+1.0 mg/L KT+MS+0.8%琼脂+3%蔗糖最适合子叶、叶柄、茎诱导愈伤,在此培养基中,茎的出愈率为96%,褐化率为0,叶柄的出愈率最高,为92%,褐化率为2%,子叶的出愈率为60%,褐化率为4%;在0.3 mg/L NAA+1.0 mg/L KT+MS+0.8%琼脂+3%蔗糖条件下最适合子叶、叶柄诱导胚性愈伤;在0.3 mg/L NAA+1.0 mg/L 6-BA+MS+0.8%琼脂+3%蔗糖条件下最适合真叶诱导胚性愈伤;在0.3 mg/L NAA+0.5 mg/L KT+MS+0.8%琼脂+3%蔗糖条件下最适合茎诱导胚性愈伤。【结论】4种外植体均产生愈伤组织,且进一步诱导愈伤组织可分化形成体细胞胚。4种外植体中,茎的愈伤诱导率最高,褐化率最低,为最佳外植体。

不同植物生长调节剂对半夏愈伤组织诱导效应的研究

目的以半夏叶柄为外植体,研究不同植物生长调节剂对半夏愈伤组织形态的影响。方法分别采用单一激素和多个激素正交试验,考察不同植物生长调节剂对半夏愈伤的诱导效果。结果单一激素不同水平的愈伤组织诱导效果因激素种类和浓度的不同而有所差异;多种激素不同水平对半夏愈伤诱导的效果与诱导率和启动时间有关,同时还应考虑愈伤的质地、形态等;植物生长调节剂对半夏愈伤组织的诱导能力从大到小依次为6-苄基腺嘌呤、α-萘乙酸、2,4-二氯苯氧乙酸、蔗糖。结论通过考察不同植物生长调节剂对半夏愈伤组织的诱导效应,确定了适宜做半夏悬浮培养材料的培养基和一次性成苗培养基。

碳源及植物生长调节剂对雷公藤愈伤组织生长和次生代谢产物含量的影响

以雷公藤根愈伤组织为材料,添加不同质量浓度的2,4-D,NAA,6-BA,KT或其组合,研究植物生长调节剂及其组合以及碳源对雷公藤愈伤组织的生长、次生代谢产物含量的影响。结果表明:2,4-D有利于雷公藤愈伤组织的生长和总生物碱的积累,6-BA对愈伤组织的生长起抑制作用,但明显促进愈伤组织中雷公藤内酯醇的形成,KT虽然对愈伤组织的生长起促进作用,但明显抑制愈伤组织中雷公藤内酯醇的形成;不同质量浓度2,4-D,NAA及与KT,6-BA组合试验表明,1.0mg·L-12,4-D+0.5mg·L-1KT明显促进愈伤组织的生长,1.0mg·L-12,4-D+2.0mg·L-1KT有利于愈伤组织中总生物碱的形成,而4.0mg·L-1NAA+0.5mg·L-16-BA有利于愈伤组织中雷公藤内酯醇的合成。碳源中,对雷公藤愈伤组织的增长量以蔗糖最高,麦芽糖最有利于雷公藤内酯醇的合成,葡萄糖最有利于雷公藤总生物碱的合成。

青钱柳愈伤组织诱导

为获得优质的愈伤组织,以青钱柳嫩茎和叶片作为外植体诱导愈伤组织,研究了不同的消毒方法、基本培养基和植物生长调节剂对青钱柳叶片和茎段愈伤组织诱导的影响。结果表明,以青钱柳茎段为外植体诱导愈伤组织,最佳的消毒组合为70%酒精10 s+0.1%升汞4 min,污染和死亡率最低(18.0%),诱导率最高(95.0%)。以青钱柳叶片为外植体诱导愈伤组织,最佳的消毒组合为70%酒精0 s+0.1%升汞4 min,污染和死亡率最低(13.7%),诱导率最高(96.3%)。以青钱柳茎段、叶片为外植体诱导愈伤组织的最佳基本培养基分别为改良MS和MS,诱导率最高分别为95.56%和96.37%。激素组合KT0.5 mg.L-1+2,4-D2.0 mg.L-1效果最佳,诱导率高达97.5%。研究结果为青钱柳愈伤组织诱导最适培养条件的筛选提供依据,同时也为进一步建立青钱柳组培快繁及悬浮培养体系提供基础。

植物生长调节剂对南方红豆杉愈伤组织培养和紫杉醇合成的影响

通过不同种类和水平植物生长调节剂对南方红豆杉(Taxus chinensisvar.mairei)愈伤组织诱导、生长和紫杉醇合成能力影响的研究发现:诱导培养初期,以无植物生长调节剂的MS为基本培养基,在附加不同植物生长调节剂组合作用下愈伤组织产生的时间和生长、在相同植物生长调节剂组合作用下不同外植体愈伤组织的产生时间和生长均表现出较显著差异,2,4-D/NAA高于0.4时,不利于南方红豆杉愈伤组织的诱导。转换到附加不同植物生长调节剂组合的B5培养基上后,随培养继代次数的增加,生长差异逐渐缩小,直至不显著,表明参考不同文献报道最优配方所设计的各植物生长调节剂组合对南方红豆杉愈伤组织的生长均较适宜,有利南方红豆杉愈伤组织生长的植物生长调节剂优化组合没有唯一性。但不同调节剂组合作用下的同源愈伤组织中、相同调节剂作用下不同源愈伤组织中紫杉醇含量均存在着极显著差异,适当水平(2 mg/L)的2,4-D单用,或与适当水平的KT、6-BA、KT+GA配合使用,对南方红豆杉愈伤组织紫杉醇的合成较有利,NAA则不太有利,幼茎和叶愈伤组织产紫杉醇的水平较其它愈伤组织为高。

不同培养基类型和植物生长调节剂配比对白桦愈伤组织中三萜积累的影响

In this study,effects of the different medium,and the kinds and concentrations of plant growth regulator on callus induction,growth and triterpenoid production in birch (Betula platyphylla) stem extant culture were investigated. The results showed that calli were all obtained on IS medium supplemented with 6-BA and NAA,KT and 2,4-D,or 6-BA and TDZ,but the shape and growth state of callus were different. The combination of 0.8 mg·L-1 6-BA+0.6 mg·L-1 NAA was the best in inducing callus from the extant,with the frequency of callus induction 100%,and the callus grew well after multiple subculture. Furthermore,the effect of 5 types of media (MS,NT,IS,WPM,B5) and different combinations of plant growth regulators on callus growth and triterpenoid accumulation were compared. It was found that WPM medium was most suitable for callus growth,the fresh weight on harvest was 6.6 g. IS medium was most suitable for triterpenoid accumulation,the content of triterpenoid on harvest was 2.37 mg·g-1 . The most beneficial combination of plant growth regulators for fresh weight was 0.4 mg·L-1 6-BA+0.1 mg·L-1 TDZ,the fresh weight on harvest was 14.08 g,while the most suitable combination of plant growth regulators for triterpenoid accumulation was 0.8 mg·L-1 6-BA+0.6 mg·L-1 NAA,the content of triterpenoid on harvest was 3.53 mg·g-1. It was concluded from above results that the best culture condition for callus growth and triterpenoid accumulation was IS+0.8 mg·L-1 6-BA+0.6 mg·L-1 NAA.

低温与植物生长调节剂预处理对小麦成熟胚培养特性的影响

为提高小麦成熟胚愈伤组织的分化率,以3个黄淮麦区冬小麦品种(系)‘小偃22’、‘西农1013’和‘千斤早’为材料,研究了4℃低温和3种植物生长调节剂(TDZ、2,4-D、多效唑)预处理小麦种子对小麦成熟胚愈伤组织分化特性的影响。结果表明:4℃低温和植物生长调节剂预处理对小麦成熟胚出愈率无显著影响,但能够显著促进愈伤组织的分化,并表现出处理及基因型间差异。‘小偃22’、‘西农1013’、‘千斤早’感应低温预处理的最好时间节点分别为6h、12h、12h,其成熟胚愈伤组织的分化率比对照分别显著提高16.2%、14.2%、12.8%;3个材料分别在10mg/L、5 mg/L、20 mg/L TDZ预处理下成熟胚愈伤组织分化率最高,比对照分别显著提高39.1%、29.7%、16.7%;3个材料的最佳2,4-D预处理浓度均为10mg/L,其愈伤组织分化率分别比对照显著提高17.3%、11.2%、25.2%;3个材料的成熟胚愈伤组织分化率分别在10mg/L、20mg/L、20mg/L多效唑预处理时最高,分别比对照显著提高6.2%、11.6%、7.2%。研究表明,低温和植物生长调节剂预处理均可有效提高小麦成熟胚愈伤组织的分化率,但促进植物生长的调节剂(TDZ和2,4-D)对小麦成熟胚愈伤组织再分化的效应远大于抑制植物生长的调节剂(多效唑)。

彩色马蹄莲叶片愈伤组织培养体系的建立

采用彩色马蹄莲(Zantedeschia spp cv Pink Giant)的叶片为外植体,运用正交设计方法,对诱导马蹄莲愈伤组织培养基进行了预试验,并对影响马蹄莲愈伤组织诱导的因素进行了定量分析;在正交试验基础上,设计了14种不同激素种类和配比的配方,进行了诱导愈伤组织体系的建立.对愈伤组织发生过程中形态学和细胞学结构进行了动态观察.结果表明:在MS基本培养基中添加6-BA和2,4-D对以叶片外植体的愈伤诱导率有显著影响;对初代诱导的愈伤组织进行的两次继代培养结果表明,继代和愈伤组织继续发生的最佳培养基为MS附加2,4-D 2.0 mg/L和BA 2.0 mg/L.

青钱柳幼胚愈伤组织的诱导

以青钱柳幼胚为外植体,研究了不同幼胚期、基本培养基种类、植物生长调节剂种类及其浓度配比对愈伤组织诱导的影响。结果表明,不同发育期的幼胚诱导出的愈伤组织在形态上差异较大,青钱柳种子发育至子叶胚中期较适合愈伤组织的诱导。青钱柳幼胚愈伤诱导时基本培养基以改良DKW效果最佳,诱导率达到了88.89%,且无褐化现象;含2,4-D与KT组合的培养基较适宜愈伤组织的诱导,其中2,4-D(0.5 mg/L)+KT(2.0 mg/L)和2,4-D(1.0 mg/L)+KT(1.0 mg/L)的组合对青钱柳幼胚诱导愈伤组织效果最佳,诱导率分别为80.00%和78.33%。

余甘子下胚轴离体再生体系的优化

以余甘子(Phyllanthus emblicaL.)下胚轴为外植体,通过器官发生途径诱导形成不定芽,探讨影响下胚轴器官发生和植株再生的因素,建立了下胚轴高效再生体系。结果表明:附加AgNO31mg·L-1和蔗糖40g·L-1的1/2MS培养基最适合不定芽的分化;下胚轴不同部位不定芽再生能力由强到弱表现为:形态学上部>中部>下部;前期暗培养有利于下胚轴不定芽的形成,暗处理14d效果最好。通过以上优化措施可以使不定芽再生率达100%,平均每外植体不定芽数11·7个。不定芽较适生根培养基为胺鲜酯(DA-6)3mg·L-1+复硝酚钠(CSN)3mg·L-1,生根率达到62·8%。