微拟球藻抗生素抗性分析和选择标记筛选

Antibiotic resistance of Nannochloropsis oculata and determination of selection markers for its genetic transformation

中文关键词: 微拟球藻  遗传转化  筛选试剂  博来霉素

英文关键词: Nannochloropsis oculata  Genetic transformation  Selection reagents  Zeocin

中文摘要:

      为了筛选到微拟球藻Nannochloropsis oculata的筛选标记，研究了微拟球藻对新霉素（Neomycin）、链霉素（Streptomycin）、潮霉素（Hygromycin）、壮观霉素（Spectinomycin）和博来霉素（Zeocin）的敏感性。每种抗生素都设置液体培养和固体培养实验并且浓度梯度相同。新霉素和链霉素的浓度梯度为：0、100、200、400、600、800 μg/ml。潮霉素的浓度梯度为0、25、50、100、125、150 μg/ml。壮观霉素浓度梯度为0、40、80、100、150、200 μg/ml。博来霉素浓度梯度则为0、0.1、0.2、0.4、0.5、1 μg/ml。结果表明，微拟球藻对新霉素、链霉素、潮霉素和壮观霉素不敏感，而对博来霉素高度敏感。在固体培养时，0.5 μg/ml博来霉素即可完全抑制微拟球藻生长；在液体培养时，1 μg/ml博来霉素即可完全抑制其生长。博来霉素可作为微拟球藻遗传转化的选择抗生素。ble基因为其阳性筛选的选择标记基因。

英文摘要:

      o determine the selection marker of N. oculata, the sensitivity to 5 antibiotics (Neomycin, Streptomycin, Hygromycin, Spectinomycin, and Zeocin) was determined in this study. For each antibiotic, liquid medium and solid medium were both adopted with the same concentration gradient. The concentration gradient of Neomycin and Streptomycin was 0, 100, 200, 400, 600, and 800μg/ml, while that of Hygromycin was 0, 25, 50, 100, 125, and 150μg/ml, and that of Spectinomycin was 0, 40, 80, 100, 150, and 200μg/ml. The concentrations of Zeocin were much lower, which were 0, 0.1, 0.2, 0.4, 0.5, and 1μg/ml. It was found that N. oculata is not sensitive to Neomycin, Streptomycin, Hygromycin or Spectinomycin, but highly sensitive to Zeocin. Zeocin can completely inhibit the growth of N. oculata when its concentration is 0.5μg/ml in solid medium or 1μg/ml in liquid medium. Zeocin could serve as a selection reagent in the genetic transformation of N. oculata, and the ble gene could be the selection marker.