원문정보
초록
영어
The combination of Simian Virus40 (SV40)’s large T antigen with its replication origin is commonly used in molecular studies to enhance the expression of heterogeneous genes through multiplying the plasmid copy number. There are no reports related to the impact of the SV40 T antigen on plant, multiple fissional, cell-type. This study explores the response of two multiple-fission microalgal cells, Scenedesmus quadricauda and Chlorella vulgaris, to the expression of the T-antigen, with aim of applying SV40 T-antigen to increase the expression efficiency of foreign genes in the two species. Different levels of low-expression have been constructed to control the expression of SV40 T antigen using three heterogenous promoters (NOS, CaMV35S, and CMV). Chlorella cultures showed slowdown in the growth rate for samples harboring the T antigen under the control of CaMV35S and CMV promoters, unlike Scenedesmus cultures which showed no significant difference between samples and could have silenced the expression.
목차
1. Introduction
2. Experiments
2.1 Isolation, purification and cultivation of Scenedesmus quadricauda and Chlorella vulgaris
2.2 Cassette construction
2.3 Vector construction
2.4 CaMV35S promoter activity
2.5 Electroporation and Cultivation
2.6 Modified protocol for screening the mitotic-division
2.7 DNA extraction, Quantitative and normal PCR
2.8 Immunofluorescence and Flow cytometry
2.9 Total RNA extraction, RT-PCR, and Transcription analysis by PCR
3. Results and discussion
3.1 Sequencing result
3.2 Polymerase chain reaction and Quantitative real time PCR
3.3 Flow cytometry analysis for Scenedesmus cultures
3.4 RT-PCR and Transcription analysis
3.5 Confirming the activity of the CaMV35S Promoter in Scenedesmus sp.
4. Conclusion
Acknowledgment
References