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- W2386813969 abstract "Objective To construct a recombinant adenovirus Ad5 / F35-HF2S with HA tag.Methods Double chains of DNA of HA tag were synthesized in vitro.FKBP1,FKBP2 and SH2 gene segments were amplified by PCR using the cDNA of K562 cells as template,and subcloned into shuttle plasmid pAdTrack-CMV-HA in turn.The constructed recombinant shuttle plasmid pAdTrack-CMV-HF2S was linearized with PmeⅠfor homologous recombination with adenoviral backbone vector pAd5 / F35 in competent Ad5 / F35-BJ5183 cells.The obtained recombinant adenovirus plasmid pAd5 / F35-HF2S was linearized with PacⅠ,and transfected to AD-293 cells for packaging and amplification.The obtained recombinant adenovirus Ad5 / F35-HF2S was subjected to two rounds of propagation and determined for titer.The transcription of HF2S mRNA and expression of HF2S protein in infected AD-293 cells were determined by PCR and Western blot respectively.Results Both recombinant shuttle plasmid pAdTrack-HF2S and recombinant adenovirus vector pAd5 / F35-HF2S were constructed correctly.After packaging and two rounds of propagation,recombinant adenovirus Ad5 / F35-HF2S reached a titer of 1013 IU / ml.PCR and Western blot proved that HF2S gene in recombinant adenovirus was successful expressed in AD-293 cells.Conclusion Recombinant adenovirus Ad5 / F35-HF2S was successfully constructed,which laid a foundation of further study on role of SH2 domain of Grb2 in gene therapy as well as the relevant mechanism." @default.
- W2386813969 created "2016-06-24" @default.
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- W2386813969 date "2012-01-01" @default.
- W2386813969 modified "2023-09-23" @default.
- W2386813969 title "Construction and identification of recombinant adenovirus Ad5/F35-HF2S" @default.
- W2386813969 hasPublicationYear "2012" @default.
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