Gadamchetty Pavan, Mullapudi Phanindra Lakshmi Venkata, Sanagala Raghavendrarao, Markandan Manickavasagam, Polumetla Ananda Kumar
1National Research Centre on Plant Biotechnology, Lal Bahadur Shastri Building, Pusa Campus, New Delhi, 110012 India.
2Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli, 620024 India.
3 Biotech. 2019 Apr;9(4):139. doi: 10.1007/s13205-019-1671-2. Epub 2019 Mar 13.
Scientific interest in microalgal species is growing and, genetic transformation has definitely opened more avenues, in the ongoing research on microphytes. In the present study, we have attempted to transform by mobilizing double-stranded linear Transfer DNA (T-DNA) comprised of () gene cassette and II (II) gene cassette non-covalently bound to TAT peptide, into cells treated with Triton X-100. The transformed cells when examined under fluorescent microscope, exhibited green fluorescence in comparison to the untransformed cells. The transformed cells were further screened, and the surviving colonies were sub-cultured, on BG11 medium fortified with Hygromycin. The surviving colonies were confirmed for the presence of integrated T-DNA by Polymerase Chain Reaction with and II gene-specific primers. This methodology has potential to substitute the existing tedious transformation methodologies and ease the future studies in microalgae.
对微藻物种的科学兴趣正在增加，并且在对微植物的 ongoing 研究中，遗传转化无疑开辟了更多途径。在本研究中，我们试图通过将由（）基因盒和非共价结合到 TAT 肽的 II（II）基因盒组成的双链线性转移 DNA（T-DNA）导入用 Triton X-100 处理的细胞中来进行转化。与未转化的细胞相比，在荧光显微镜下检查时，转化后的细胞呈现绿色荧光。对转化后的细胞进行进一步筛选，并将存活的菌落进行传代培养，置于添加潮霉素的 BG11 培养基上。通过使用和 II 基因特异性引物的聚合酶链反应确认存活菌落中是否存在整合的 T-DNA。这种方法有潜力替代现有的繁琐转化方法，并简化未来微藻研究。
