荸荠组培苗在间歇浸没式生物反应器（TIBs）中的高效增殖技术

摘要

【目的】探讨间歇浸没式生物反应器（TIBs）对荸荠（Eleocharis dulcis）组培苗快速繁殖的效果，为提高荸荠组培苗繁殖效率及其工厂化生产提供技术支撑。【方法】以初代诱导培养获得的桂蹄2号荸荠组培苗为外植体，利用TIBs系统进行组培快繁，研究荸荠组培苗不同继代代数、接种密度、激素组合及反应器间歇浸没频率等因子对荸荠组培增殖的影响。【结果】经初代诱导培养的第4、5、6代继代材料增殖倍数较高，其中第6代继代苗增殖倍数最高，达31.1倍，到第7代后随继代代数的增加增殖倍数逐渐降低。当接种密度大于15丛/瓶时，外植体污染率达16.0%；当接种密度低于10丛/瓶时，外植体的污染率为0。低于10 min/6 h的间歇浸没频率有利于降低污染率，在6 h内低于15 min的浸没时间有利于降低玻璃化苗率；当激素组合为3.0 mg/L 6-BA+0.01 mg/L NAA时，组培苗增殖倍数达38.5倍，且组培苗生长正常。【结论】在TIBs系统中，采用经初代诱导培养的第5代荸荠继代组培苗，增殖培养基为MS+3.0 mg/L 6-BA+0.01 mg/L NAA+30.0 g/L蔗糖，pH 6.0，接种密度10丛/瓶，反应器间歇浸没频率为10 min/6 h，可获得较好的培养效果。%Objective]Rapid propagation of Eleocharis dulcis plantlets in temporary immersion bioreactors (TIBs) was explored to improve its propagation efficiency , and provide technical support for E. dulcis plantlets factory production.[Method]E. dulcis variety Guiti 2 plantlets which obtained through primary inducement culture were taken as explants and propagated using TIBs. Influence of factors such as different subcultures, inoculation densities, different combinations of hormones and different immersion frequency on E. dulcis plantlets propagation were studied. [Result]In TIBs system, multiplication rates of the fourth, fifth and sixth subcultures were high. The sixth subculture had the highest multiplication rate, reaching 31.1 times. The multiplication rates declined after the seventh subculture. When inoculation density was more than 15 clusters per bottle, explant contamination rate reached 16.0%; whereas when the inoculation density was less than 10 clusters per bottle, contamination rate was 0%. When the intermittent immersion frequency was less than 10 min/6 h was beneficial for reducing contamination rate. Less than 15 min/6 h immersion time was beneficial for reducing vitrification seedling rate. With 6-BA 3.0 mg/L+NAA 0.01 mg/L combination of hormones , multiplication rate of the first plantlets was 38.5 times, and the plants were in normal growth. [Conclusion]In TIBs system, the fifth subculture with MS+6-BA 3.0 mg/L+NAA 0.01 mg/L+sugar 30.0 g/L as medium, pH value being 6.0, inoculation destiny being ten clus-ter per bottle and immersion frequency being 10 min/6 h can achieve considerable cultivation effect.