香蕉组织培养过程中内生菌污染的控制

Control of Endogenous Bacterial Contamination and Micropropagation of a Traditional Table Banana (Musa spp. cv. Kanthali) of Bangladesh

摘要:

研究了体外培养一种孟加拉传统香蕉（Musa spp. Cv. Kanthali）的茎尖组织。茎尖的原始细胞表面经无菌处理（0.1% HgCl₂处理12min），接种6～15d后外植体地下茎部分仍有微生物污染（大部分是细菌），杀死了85％的外植体。为确定无污染培养基，将等量外植体分别浸泡在含400mg/L氨苄青霉素和200mg/L庆大霉素（两种光谱抗生素）的培养基中1h。结果表明，经抗生素处理的外植体完全没有污染，但培养3周后不能再生。进行二次继代培养后，其中一部分外植体吸收了培养基并胀大，颜色由苍白转变成浅绿或深绿。三次继代培养后数天，不再观察到外植体的生长，所有经抗生素处理过的外植体都开始死亡。在未经抗生素处理的活外植体中，单个茎发育的最佳培养基是：MS+4.0mg/L BA+0.5mg/L KT+15% CW，平均生长时间为18～21d，但再生率很低，只有30％。茎细胞增殖的最佳培养基是：MS+4.0mg/L BA+2.0mg/L IAA+15% CW，每个茎平均只萌发3～4个芽。最后，在添加0.5mg/L IBA的一半浓度的MS培养基中，体外培养茎最大生根率达到90％。

Abstract:

Shoot tips of a traditional table banana (Musa spp. cv. Kanthali) of Bangladesh were evaluated for in vitro propagation. Initial surface sterilization (with 0.1% HgCl₂ for 12 minutes) of shoot tips was successful but microbial contamination (mostly bacteria) at the rhizomatous base of the explants was observed within 6～15 days after inoculation which eventually killed 85% of inoculated explants. So, for contamination free culture establishment explants were soaked in two broad spectrum antibiotics namely ampicillin and gentamicin. Cent percent contamination free cultures were established by soaking the explants in 400mg/L ampicillin or 200mg/L gentamicin for 1h. Antibiotic treated explants were found to be full contamination free but failed to regenerate after 3 weeks of culture. But some of them absorbed media for up to 2^nd subculture and showed swelling of explants and some color changes from pale white to light/deep green. Finally, a few days after 3^rd subculture, no growth of explants was observed and all treated explants eventually started to die. Among the untreated alive explants the best medium for single shoot development was MS+4.0mg/L BA+0.5mg/L KT+15% CW and average time required for shoot development was 18～21 days. But the regeneration percentage was very low (30%). The best medium for shoot multiplication was MS+4.0mg/L BA+2.0mg/L IAA+15% CW and only average 3～4 shoots were formed per shoot. Finally, in vitro proliferated shoots produced roots with maximum frequency (90%) in half strength of MS medium fortified with 0.5mg/L IBA.