条斑紫菜壳孢子的抗生素敏感性研究

Studies on antibiotic sensitivity of conchospores in Porphyra yezoensis Ueda(Bangiales,Rhodophyta)

以往研究紫菜细胞对抗生素的敏感性时,常以叶状体的酶解单离体细胞为材料,但由于来自同一个叶状体的体细胞处于不同的分化阶段,离体培养时,体细胞的成活、再生和发育途径均不相同,从而无法准确地判断抗生素对紫菜离体细胞的生长和发育影响。以成活、萌发和发育途径等几乎一致的条斑紫菜壳孢子为研究材料,较系统地观察了3种抗生素对壳孢子的成活、生长和发育的影响。结果表明:卡那霉素能促进条斑紫菜壳孢子的萌发、生长和发育。加卡那霉素培养7d,26%的壳孢子萌发体,其细胞数多于20个,比对照组多2倍,培养11d和18d,萌发体的平均长度分别为对照组的1.6倍和2.1倍。氨苄青霉素能显著地抑制条斑紫菜的壳孢子萌发、生长和发育,造成细胞的分裂速度减缓、叶状体发育异常。加氨苄青霉素培养7d,80%的壳孢子萌发体的细胞数少于15个,培养11d和18d,各试验组的壳孢子萌发体的平均长度均显著低于对照组,并且随着氨苄青霉素浓度的提高,萌发体的平均长度下降。氯霉素对条斑紫菜的壳孢子具有强烈的致死作用。加氯霉素培养7d,浓度大于100μg/mL的各试验组的壳孢子萌发体全部死亡;培养11d,低浓度组(50μg/mL)的萌发体也全部死亡,90%以上的萌发体的细胞数小于5个。由此得出,氯霉素是开展条斑紫菜基因工程的理想抗性选择压力。

In the previous studies on the antibiotic sensitivity of Porphyra,the somatic cells isolated enzymatically from Porphyra gametophytic blades were often used.However,the accurate analysis of antibiotics effects on the survival,growth and development of the somatic cells was difficult,because the somatic cells isolated even from a Porphyra blade had different regeneration capacity,cell division and developmental pathways when they were cultured in vitro.In this paper,the antibiotic sensitivities of Porphyra yezoensis conchospores which are almost the same in viability,germinability and developmental pathways,were studied by treatment with kanamycin,ampicillin and chloramphenicol.The obtained results demonstrated that kanamycin could promote the growth and development of conchospore germlings.After 7 days of culture,the cell number of 26% of the conchospore germlings was over 20 cells,which was twice as many as that of control group.After 11 and 18 days of culture,the mean length of conchospore germlings was 1.6 times and 2.1 times longer than that of control group,respectively.The treatment with ampicillin could inhibit the growth and development of conchospore germlings,showing decrease of cell division rates and inhibitation of germling development.After 7 days of culture,the cell number was less than 15 in 80% of conchospore germlings.After 11 and 18 days of culture,the mean length of conchospore germlings was shorter than that of control group,as ampicillin concentrations increased.The chloramphenicol had strong lethal effect on conchospore germlings.When chloramphenicol concentration was higher than 100 μg/mL,the conchospore germlings died out after 7 days of culture.The conchospore germlings treated with 50 μg/mL of chloramphenicol also died out after 11 days of culture.The cell number was less than 5 cells in 90% of the dead conchospore germlings.Therefore,it was concluded that the chloramphenicol is an effective selective pressure in genetic engineering of P.yezoensis.