An Approach for Micropropagation of Blueberry (<i>Vaccinium corymbosum</i>L.) Plants Mediated by Temporary Immersion Bioreactors (TIBs)

A new procedure for blueberry (Vaccinium corymbosum L.) micropropagation in programmed Temporary Immersion Bioreactors (TIBs based on two separate bottles) was developed for the commercial genotypes Biloxi, Sharp Blue and Brillita. Plant cultures were developed in a controlled environment with 0.4 MPa CO2 enrichment, sucrose-reduced medium, and light intensity of 60 mM m-2·s-1. Principal component analysis showed that component 1 (C1) grouped 64.08% of the total variability, while the first two components accounted for 86.97%. Representation of the principal components demonstrated three clusters corresponding with the blueberry genotypes, and within each cluster plants micropropagated in agar-base medium grouped separately from those plants multiplied in TIBs. Both plant number and total internodes traits (related to the productive efficiency) were demonstrated superior in blueberries propagated in TIBs. Additionally, when transferred to greenhouse conditions, blueberries propagated in TIBs showed higher adaptability and growing rates than those cultured by the conventional approach, altogether evidencing the occurrence of a photomixotrophic stage in the vitroplantlets cultured in TIBs.

Design and Construction of a Pneumatic Temporary Immersion Bioreactor System for the Multiplication of <i>Ananas comosus</i>var. Trujillana Red

The present work initially identified the design parameters of a temporary immersion bioreactor to later scale it to a complete system for the in vitro multiplication of Ananas comosus var. Trujillana Red. Thus, a low-cost pneumatic temporary immersion bioreactor system was designed and built with 24 tanks of 2 L each. The automation of the system was designed and implemented by means of a timer circuit whose design parameters were: duration of the propagation process, which depends on the multiplication period of the crop and is an open variable, which means that the operator decides when to turn off the system;the duration of each dive, which for reasons of complexity of the algorithm was standardized as one minute;immersion frequency, which was programmed for intervals of 1, 2, 3, 4, 5, 6, 7, 8 hours respectively and duration of aeration, which from a test run times of 0.20 were chosen, 30, 40, 50, 60, 70, and 80 seconds that correspond to the time of delivery of compressed air;additionally, the multiplication rate of Ananas comosus var. Trujillana Red in the immersion system which was 6.5 times per propagative unit inoculated in thirty days.

High Efficiency of Propagation for Sagittaria sagittifolia Using a Temporary Immersion Bioreactor System

Although several studies have reported on the propagation of the Sagittaria sagittifolia to date, none of these methods have efficiently achieved the mass production of these plants. The present study aimed to investigate the propagation and growth of S. sagittifolia using a temporary immersion bioreactor system （TIBS） compared with conventional semi-solid and liquid culture. The effect of different immersion frequencies and immersion times together with supplementation of various plant growth regulators to the Murashige and Skoog （MS） medium was evaluated on shoot proliferation and plant growth. The results showed that the higher immersion frequency （every 6 h） and shorter immersion time （3 min and 10 rain） in medium containing 4 mg/L BA and 0.1 mg/L NAA produced the highest multiplication rate （23）, which are significantly higher than conventional semi-solid （3.6） and liquid （4.5） method, and the best plant growth parameter. While, the lower immersion frequency and longer immersion time （30 rain every 12 h and 60 min every 24 h） induced vitrification and pollution rate in shoot tips 16.6% and 19%, 42% and 37%, respectively. There is distinct decrease in pollution rate （8.3%） in TIBS （10 min every 6 h） compared with the conventional semi-solid and liquid cultures. Medium containing 4 mg/L BA and 0.5 mg/L NAA using 10 min immersion every 6 h showed satisfaction at the rooting stage, with high shoot proliferation rate （21.6）, 100% rooting and 94% plant survival. Therefore, applying TIBS in S. sagittifolia is an efficient method for scaling up the production of plantlets with high quality seedlings.

Rapid Propagation of Virus-free Sugarcane Plantlets via Temporary Immersion Bioreactor System

By employing temporary immersion bioreactor system(TIBs),we studied virus-free culture of seedlings from sugarcane varieties ROC16 and ROC22,from medium recipe,inoculation amount,sucrose concentration,and variety difference. The results showed,using this method,that proliferation rate of ROC16 improved by 40 times,per flask generated about 800 plantlets; of ROC22 improved by 30 times,per flask generated about 400-600 plantlets. The results provided basis for using TIBs in rapid propagation of plantlets via tissue culture.

TIBs和传统培养方式培养的甘蔗脱毒苗生理生化指标比较

以新型培养方式——间歇浸没式生物反应器(TIBs)和传统培养方式(CK)生产的甘蔗脱毒苗为材料,分别在增殖期、生根期和炼苗期测定它们的生理生化指标并进行比较。结果表明,在株型方面,TIBs脱毒苗在株高、茎径、叶片数、叶长、叶宽等方面均比CK的高;在含水量方面,TIBs和CK的脱毒苗差别不大;在叶绿素、脯氨酸、可溶性糖、可溶性蛋白含量和过氧化物酶活性等指标方面,TIBs与CK的脱毒苗相比在增殖期差异不大,但到了生根期和炼苗期各指标均比CK的高。应用间歇浸没式生物反应器培养的甘蔗脱毒苗比传统培养方式具有更强的生长活力和抗逆性。

Simple Protocol for the Micropropagation of Teak(Tectona grandis Linn.)in Semi-Solid and Liquid Media in RITA^(█) Bioreactors and ex Vitro Rooting

In Latin America the forestry of exotic species such as teak has been increasing in recent decades, due to their advantages in wood quality, rapid growth;and the relative ease of producing clones and their multiplication with respect to native species. Therefore, there is great interest in developing larger-scale propagation strategies that reduce costs and intensive manual labor. Culture in liquid media with temporary immersion and the semi-automation of the system has raised expectations for large-scale micropropagation. We report a protocol for teak, which reuses the primary explants in several culture cycles in semi-solid medium to produce nodal explants for the multiplication phase in temporary immersion bioreactors (RITA&reg;). The control of factors such as cytokinin concentration, explants density, immersion frequencies and culture duration was analyzed. The number of shoots increased with 0.5 mg&middot;l-1 of BA (6-Benzyladenine), alone or in combination with 0.5 mg&middot;l-1 of Kinetin, with 2 daily immersions of 1 minute each;however, these shoots showed a high degree of hyperhydricity. When 0.05 mg&middot;l-1 of BA was used with 1 immersion of 1 minute every 2 days, the hyperhydricity decreased. Although the number of shoots was lower, they showed good length to be used during multiplication and rooting ex vitro. Our results suggest that teak micropropagation can be simplified in two phases in vitro, the establishment and multiplication;followed by rooting ex vitro and acclimatization. This would imply a reduction in production costs, since most of the multiplication would take place in RITA&reg;containers.

TIB for Micropropagation and the Relationship between Anthocyanins and Chlorophyll of Strawberry Seedlings

In this study, we employed a temporary immersion bioreactor(TIB)system for the micropropagation of strawberry seedlings. The TIB method and the conventional method for the micropropagation of strawberry seedlings were compared in terms of explant propagation coefficient, seedling fresh weight,contents of anthocyanins and chlorophyll, and photosynthetic characteristics. The results showed that an inoculation density of 40 explants/L was suitable for the micropropagation of ’Benihope’ strawberry seedlings in the TIB. The propagation coefficient, fresh weight, photosynthetic rate, stomatal conductance, transpiration coefficient, and the contents of total anthocyanins and chlorophyll of seedlings micropropagated in the TIB were significantly higher than those of the seedlings micropropagated by the conventional method. In conclusion, the TIB method was superior to the conventional method in the micropropagation of strawberry seedlings.The negative reciprocity did not occur between anthocyanins and chlorophyll in strawberry seedlings cultured under forced ventilation such as TIB.

利用间歇浸没式生物反应器进行甘蔗组培快繁的研究

以甘蔗(新台糖22号)茎尖脱毒组培苗为材料,利用间歇浸没式生物反应器(TIBs)进行组培快繁的技术体系研究.结果表明:TIBs系统可使甘蔗组织培养一代增殖40倍以上,远远高于传统方法;TIBs系统以第3代继代的材料为宜;10～15株/瓶的接种密度最有利于组培苗的增殖;激素选择与传统方法相似,0.5～1.0 mg/L 6-BA适合甘蔗组织的增殖,4 mg/L NAA有利于根的诱导;浸没间歇频率以浸没1 min间歇3 h处理的增殖和生长表现比较优异,而6 h时则有利于根的诱导和根的生长.

基于间歇浸没式生物反应器的荔浦芋组培快繁体系优化

【目的】优化基于间歇浸没式生物反应器系统(TIBs)的荔浦芋组培快繁体系,为荔浦芋脱毒组培苗的工厂化、自动化生产提供技术支持。【方法】以荔浦芋新品种桂芋2号茎尖分生组织脱毒诱导获得的不定芽为外植体,利用TIBs培养系统筛选出适合组培苗增殖及生根的最佳激素组合,并分析不同继代材料、接种密度及浸没间歇频率对组培苗增殖和生根效果的影响。【结果】利用TIBs培养系统可有效提高荔浦芋组培苗的增殖效果,增殖倍数达28.96倍,约是传统固体培养方法(2.87倍)的10倍,且组培苗的株高和生根数均极显著高于传统固体培养植株(P<0.01)。在TIBs培养系统中,含4.00 mg/L 6-苄氨基嘌呤(6-BA)+0.05 mg/L萘乙酸(NAA)的培养基最适合荔浦芋组培苗增殖和生长,其组培苗增殖倍数为32.04倍。当接种材料为第4代荔浦芋继代材料、接种密度为10株/L、浸没间歇频率为5 min/6 h时,最有利于组培苗增殖和生长,增殖倍数均在30.00倍以上,可缩短萌芽时间,组培苗长势良好,且培养基污染率为0。【结论】利用TIBs培养系统对荔浦芋继代材料进行高效快繁具有可行性,可为实现及推动荔浦芋健康种苗繁育的工厂化生产提供技术支持。

利用间歇浸没式生物反应器进行果蔗组培快繁

以果蔗(Badila)茎尖脱毒组培苗为材料,利用间歇浸没式生物反应器(Temporary immersion bioreactors TIBs)进行组培快繁的技术体系的研究。结果表明:(1)使用TIBs系统进行甘蔗组织培养一代增殖较传统方法高出10倍以上;(2)TIBs系统以第4代继代材料为宜;(3)接种密度在10～15株/L培养基最有利于甘蔗组培苗的增殖和生长;(4)TIBs系统中6-BA浓度为0.5～1.0mg/L时适合于增殖培养,NAA浓度为4mg/L时有利于根的诱导;(5)浸没间歇频率在1min/3h时增殖和生长表现较为优异,1min/6h时有利于根的诱导和根的生长。

何首乌离体快繁技术体系的建立

【目的】建立何首乌离体快繁技术体系,为其种苗生产提供技术支持。【方法】以从广西陆川县采集的何首乌幼嫩单芽茎段为外植体,分析不同灭菌时间、不同植物生长调节剂种类和浓度及不同培养方式对其组织培养快繁技术的影响。【结果】适宜何首乌外植体消毒灭菌的方法为0.1%HgCl2浸泡处理8 min;适宜不定芽诱导的初代培养基为MS+1.0 mg/L 6-氨基腺嘌呤(6-BA)+0.05 mg/L a-萘乙酸(NAA),腋芽萌发率达96.7%;适宜的继代增殖培养基为MS+2.0 mg/L苯基噻二唑脲(TDZ)+0.05 mg/L NAA,增殖系数为5.68,植株健壮;无菌苗的单芽茎段以浸没间歇式培养(TIBs)增殖系统效果最佳;适宜的生根培养基为1/2MS+0.01 mg/L NAA,生根率达91.33%。【结论】建立了以带腋芽茎段为外植体的何首乌离体快繁技术体系,可为其种苗生产提供新途径。

利用间歇浸没式生物反应器进行慈姑组培快繁研究

以慈姑(Sagittaria sagittifolia)茎尖诱导的组培苗为材料,利用间歇浸没式生物反应器(TIBs)开展慈姑组培快繁激素组合的筛选、慈姑不同代数继代材料的培养效果、不同接种密度对慈姑组培增殖影响及不同间歇频率对慈姑组培快繁的影响等组培快繁技术体系研究。结果表明:TIBs系统可以使慈姑组培苗一代增殖19.5倍以上,比传统方法提高3倍以上;培养基含6-BA 3.0mg/L+NAA 0.01 mg/L的TIBs适合组培苗继代增殖培育;TIBs系统中,以第5代继代材料为宜,接种密度10株/L,间歇浸没频率在10 min/3 h时,最有利于慈姑组培苗的增殖和生长。

不同组培方法对香蕉组培苗假植阶段生理特征的影响

以培养获得的第6代香蕉不定芽为试验材料,以两种组培方式(间歇浸没式生物反应器TIBs、传统半固体培养基)培养香蕉组培苗,进行假植移栽后20、40、60 d,分别测定不同组培苗叶片的光合特性及抗氧化酶活性等指标,比较两种组培方式对香蕉组培苗假植阶段的光合生理的影响,分析TIBs系统培养的香蕉组培苗对自然环境的适应能力。结果表明,随着移栽天数的增加,两种香蕉组培苗叶片光合速率、气孔导度等逐渐增加,而胞间CO2浓度逐渐下降,其中以TIBs培养的香蕉组培苗表现较高;两种香蕉组培苗叶片叶绿素含量及SOD、POD活性均随移栽天数的延长而上升,均以TIBs方式培养的组培苗较高;叶片MDA含量随移栽时间的延长而降低,以传统培养苗表现较高。说明在移栽后60 d内,TIBs培养的香蕉组培苗光合作用、养分吸收能力及抗逆能力均优于传统半固体培养的组培苗,TIBs可作为香蕉组培苗快繁的有效方法。

间歇浸没式生物反应器在大规模组织培养中的应用研究

介绍了组织培养中4种间歇浸没式生物反应器,即叶轮驱动和蠕动泵驱动的间歇浸没式反应器及2种气体驱动的间歇浸没式生物反应器及其研究进展;综述了农林业中实现商业化生产和正在研究的一些树种以及存在的问题。

间歇浸没式生物反应器快繁草莓脱毒苗的参数

以茎尖脱毒处理诱导的草莓‘红颜’第3代继代苗为试验材料,利用间歇浸没式生物反应器进行组培苗增殖技术体系的研究。经过40d的增殖培养后,结果表明:(1)利用TIBs系统进行草莓组培苗培养增殖率约是传统培养方式的3倍;(2)TIBs系统中接种密度在2株/0.25L培养基/瓶,有利于草莓组培苗的生长;(3)TDZ和NAA质量浓度均为0.5mg/L时有利于TIBs系统中草莓组培苗的增殖;(4)TIBs系统中蔗糖质量浓度为30g/L较适宜;(5)荧光灯与红蓝LED灯对TIBs系统中草莓增殖无差异。

利用间歇浸没植物生物反应器进行半夏组培快繁的研究

研究了一种自主开发的间歇浸没式植物生物反应器进行三叶半夏(Pinellia ternata(Thunb.)Breit.)的组培快繁。首先考察了反应器的浸没频率对半夏培养的影响,利用正交试验设计了9组不同的浸没频率,经过8周的培养,对增殖倍数、生物量、组培苗外观形态进行考察,确定反应器培养半夏的最优浸没频率为5 min/12 h;其次,在反应器连续培养的过程中,检测培养液中pH值的变化,发现第4周是节点,在此之前pH逐渐降低,之后逐渐升高,说明营养消耗和代谢产物积累主导地位的变化发生在第4周,也可以说明在培养的第4周更换新鲜的培养基或进行培养基添加能够更好的培养半夏组培苗;最后,比较了反应器和固体培养在半夏组培快繁中的特点,通过对增殖情况、外观形态和组培苗活性成分含量考量,发现反应器培养的组培苗更加健壮。显示在半夏的组培快繁中能够即时换气和间歇浸没的液体培养方式比固体培养更有优势。本文为半夏的组培快繁提供一个新的技术和方法,为半夏和其它药用植物组培苗的规模化扩繁提供了借鉴。根据实验结果,对间歇浸没式植物生物反应器在种苗扩繁、抗性(耐盐)品种筛选和次生代谢产物研究方面的用途,进行了分析讨论。

荸荠不同组培快繁方式的对比研究

以广西农科院生物所选育的荸荠组培苗桂蹄2号为材料,利用间歇浸没式生物反应器(TIBs)与传统固体组培进行快繁培养,对比研究两种技术体系。研究结果表明,TIBs系统可使荸荠组织培养一代增殖40倍以上,远远高于传统方法,且培养出的植株分蘖多,叶状茎粗,生根少,长势好。

不同组培技术对菊花生长的影响

[目的]对比间歇浸没培养技术和传统固体培养技术的优劣。[方法]分别采用间歇浸没培养技术和传统的固体培养技术进行菊花的继代培养。[结果]液体间歇浸没培养下,菊花的各项生长指标如株高、茎段数、干鲜重等均优于传统固体培养;液体间歇浸没培养下菊花的叶片水势比固体培养的低,但没有玻璃化现象发生;液体间歇浸没培养下,继代和生根过程合二为一,缩短了培养时间;在液体间歇浸没培养下,气孔面积是固体培养下的2.1倍,气孔频度是固体培养的1.8倍。[结论]间歇浸没培养方式相对于传统的固体培养在各项生理指标上表现较好,可用于大规模培养菊花。

间歇浸没式生物反应器在植物组织培养中的应用研究进展

间歇浸没式生物反应器是植物组织培养中的一种新兴技术装备,其基本原理是在无菌条件下将组培苗在培养液中进行周期性浸泡培养以实现快速繁殖。与传统组织培养相比,间歇浸没式生物反应器改进了液体培养的方式,在这种培养方式下,植物组织与液体培养基紧密接触,可刺激和促进植物组织对营养元素和激素的吸收;间歇及连续振动给液体培养基提供了足够氧气,提高了组培苗的繁殖系数。间歇浸没式生物反应器主要应用于快繁后期繁殖苗和生根苗的培育,较好地促进了自动化、机械化和商业化的植物组培快繁技术的发展。因此,间歇浸没式生物反应器具有自动化程度高、生产成本低、组培苗质量好、繁殖系数高、移栽易成活及适应性较强等特点,已在多种植物组织培养上得到应用。为更好地将该系统应用于植物组织培养,文章概述了间歇浸没式生物反应器的发展、参数设置、快繁的优势和缺点等方面的进展情况;同时对间歇浸没式生物反应器技术在植物组织培养中的研究进行展望。

利用间歇浸没式生物反应器进行甘蔗脱毒苗快繁研究

利用新型间歇浸没式生物反应系统(TIBs)在培养基配方、接种数量、蔗糖浓度及品种差异等方面对甘蔗脱毒组培苗快繁进行研究。结果表明:①PP333及6-BA+PP333均能促进组培苗分化,增殖率较高且生长整齐,可进入下一阶段培养;②GA3浓度为1.0mg·L-1伸长培养效果最好,平均苗高9.87cm;③每升培养液接种量为20株时的增殖率最高;④培养基蔗糖浓度以20g·L-1较为适宜。利用该系统对ROC16和ROC22两个甘蔗品种的脱毒苗经过3个阶段的培养,ROC16增殖率达40倍,每瓶得苗800株苗左右;ROC22增殖率为30倍,每瓶苗的数量达600株。