Construction and Identification of Eukaryotic Expression Plasmid PcDNA3.1-CSRP2-HA

Objective: To construct and identify eukaryotic expression plasmid PcDNA3.1-CSRP2-HA. Methods: According to the sequence of CSRP2 CDS in GeneBank, a Pair of Primers were respectively designed and Synthesized, and the total RNA was isolated from A549 cells. After amplification with reverse transcription polymerase chain reaction(RT-PCR), the product was cloned into PcDNA3.1-HA vector, and they were identified by PCR and double restrictive edonuclease digestion and sequence analysis. Then the recombinant expression plasmid was transferred into A549 cells, and the CRP2 protein expression was identified by Western-blot.Results: The PcDNA3.1-CSRP2-HA eukaryotic expression plasmid was successfully established, and the expression of PcDNA3.1-CSRP2-HA could be detected in A549 by Western-blot. Conclusions: The PcDNA3.1-CSRP2-HA eukaryotic expression plasmid was constructed successfully, which lays the foundation for the research of CRP2 in transcription regulation mechanism of oxidative damage induced by inflammation.