@article{Li2026, 
author = {Wenjin Li and Yilin Han and Guangdong Ji and Yanxin Hu and Pingping Wang and Zhenhui Liu},
title = {Optimization of Electroporation Conditions and Screening of Nitrate Reductase Gene Mutants in Dunaliella viridis},
year = {2026},
journal = {Periodical of Ocean University of China},
volume = {56},
number = {9},
pages = {56-66},
keywords = {Dunaliella viridis, genetic transformation, NR-deficient mutant algal, selective marker, transgene},
url = {https://www.sciopen.com/article/10.16441/j.cnki.hdxb.20250197},
doi = {10.16441/j.cnki.hdxb.20250197},
abstract = {To achieve efficient genetic transformation in Dunaliella viridis, this study optimized key parameters including culture media, antibiotic selection markers, and electroporation conditions. The results demonstrated that adding 0.5% agar to solid media enabled normal algal growth. The D. viridis strain showed insensitivity to glufosinate-ammonium but was susceptible to chloramphenicol and chlorate, with growth significantly inhibited by 400 μg/mL chloramphenicol in liquid media and 300 μg/mL in solid plates. For electroporation, cells were harvested by centrifugation at 1 700g for 3 min to achieve optimal cell collection efficiency. Using the NR gene knockout vector pKS-diaCas9-sgRNA-NRKO delivered via electroporation, nitrate reductase (NR)-deficient mutants were successfully selected on 100 mmol/L (30 g/L) sodium chlorate plate. Sequencing confirmed precise knockout at the second exon of the NR gene. This work establishes a foundational protocol for employing D. viridis as a bioreactor in genetic engineering applications.}
}