Extraction and reverse transcription of total rna from mouse brain-derived endothelial cells.3 infected by streptococcus Suis 2
Abstract
Streptococcus suis 2 is an important emerging zoonotic pathogen. It mainly causes meningitis in pigs. We use SS2 to infect bEnd.3 to get stable cDNA for next research on differences in gene expression and protein expression of cytokines.
The paper presents an SS2 study for bEnd.3 infection to obtain stable cDNA for subsequent study of differences in gene expression and cytokine protein expression.
Objective: The aim of this study was to extract the total RNA from mouse brain-derived Endothelial cells (bEnd.3) infected by Streptococcus suis serotype 2 (SS2) and transcript to complementary DNA (cDNA).
Materials and methods: SS2 strain were obtained from Jilin University, China. BEnd.3 was from Henan institute of Science of Technology, China. Reverse transcription kit was from Takara company, Japan. Trizol was from Bioteke company,China. Nanodrop instrument was from Thermo company, USA. Polymerase chain reaction (PCR) instrument was from Biometra company, Germany.
We used SS2 to infect bEnd.3 at a multiplicity of infection (MOI) of 100 for 12h. Cells were harvested and Trizol method was chose to extract the total RNA of bEnd.3 infected by SS2. Nanodrop instrument was used to measure the concentration of RNA and the values of OD260/280 and OD260/230. RNA were transcripted to cDNA with reverse transcription kit by PCR instrument.
Results: trizol method used in this study was reliable and high-quality RNA were obtained. Stable cDNA were obtained by reverse transcription kit.
Conclusion: in this experiment high-quality RNA was obtained and reverse transcribed to stable cDNA for subsequent detection of related cytokines. This study provides an approximate RNA extraction method and good experimental foundation for downstream research.
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References
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Copyright (c) 2021 Mingcheng Liu, Oksana Kasianenko
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