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High efficiency transformation by electroporation of the freshwater alga Nannochloropsis limnetica

  • Yiwen Chen
  • Hanhua HuEmail author
Original Paper
  • 105 Downloads

Abstract

The microalgal genus of Nannochloropsis is considered one of the most promising organisms for the production of biofuels due to their high lipid content. Transformation systems for marine Nannochloropsis species have been established in the recent decade, however, genetic manipulation of Nannochloropsis limnetica, the only known freshwater species in this genus, is not yet available. Based on established marine Nannochloropsis species electrotransformation protocol, nuclear genetic transformation was established in N. limnetica, meanwhile the appropriate antibiotic selection concentration and electric field strength of electroporation were determined. For the selection of transformants in N. limnetica on plates, 0.07 μg mL−1 of zeocin or 5 μg mL−1 of hygromycin B was proved sufficient, and the transformation efficiency was < 2 × 10−8 with a single pulse ranging from 2200 to 2600 V using 2-mm electroporation cuvettes. Pretreatment of N. limnetica with 10 mM lithium acetate and 3 mM dithiothreitol before electroporation increased transformation efficiency hundreds of times, and the highest transformation efficiency of 10–11 × 10−6 was obtained with an electric field strength of 12,000 V/cm. Our results help to expand the biotechnological applications of this freshwater species and provide means for successful electrotransformation of other microalgae as well.

Graphic abstract

High-efficiency transformation of freshwater Nannochloropsis pretreatment of N. limnetica with 10 mM lithium acetate and 3 mM dithiothreitol before electroporation increased transformation efficiency hundreds of times.

Keywords

Biofuels Electroporation Freshwater microalgae Nannochloropsis limnetica Nuclear transformation 

Notes

Acknowledgements

This work was supported by the National Key R&D Program of China (Grant No. 2018YFD0901500) and National Natural Science Foundation of China (Grant No. 91751117).

Author contributions

HH, taking responsibility for the integrity of the work as a whole, designed the research, analyzed the data and wrote the manuscript. YC performed the experiments and analyze the data. All authors agree on the authorship and submission of the manuscript for peer review.

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Informed consent

No informed consent, human or animal rights applicable.

Supplementary material

11274_2019_2695_MOESM1_ESM.doc (37 kb)
Supplementary file1 (DOC 37 kb)

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Key Laboratory of Algal Biology, Institute of HydrobiologyChinese Academy of SciencesWuhanChina
  2. 2.University of Chinese Academy of SciencesBeijingChina

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