Sprecher Brittany N, Zhang Huan, Lin Senjie
Department of Marine Sciences, University of Connecticut, 1080 Shennecossett Rd, Groton, CT 06340, USA.
Microorganisms. 2020 Jan 16;8(1):126. doi: 10.3390/microorganisms8010126.
The lack of a robust gene transformation tool that allows proper expression of foreign genes and functional testing for the vast number of nuclear genes in dinoflagellates has greatly hampered our understanding of the fundamental biology in this ecologically important and evolutionarily unique lineage of microeukaryotes. Here, we report the development of a dinoflagellate expression vector containing various DNA elements from phylogenetically separate dinoflagellate lineages, an electroporation protocol, and successful expression of introduced genes in an early branching dinoflagellate, . This protocol, involving the use of Lonza's Nucleofector and a codon-optimized antibiotic resistance gene, has been successfully used to produce consistent results in several independent experiments for . It is anticipated that this protocol will be adaptable for other dinoflagellates and will allow characterization of many novel dinoflagellate genes.
缺乏一种强大的基因转化工具,无法使外源基因在甲藻中正确表达并对大量核基因进行功能测试,这极大地阻碍了我们对这种在生态上重要且在进化上独特的微真核生物谱系的基本生物学的理解。在此,我们报告了一种甲藻表达载体的开发,该载体包含来自系统发育上不同的甲藻谱系的各种DNA元件、一种电穿孔方案,以及在早期分支的甲藻中成功表达导入的基因。该方案涉及使用Lonza的细胞核转染仪和密码子优化的抗生素抗性基因,已成功用于在多个独立实验中为……产生一致的结果。预计该方案将适用于其他甲藻,并将有助于对许多新的甲藻基因进行表征。