Cloning of rat serum amyloid P component gene and preparation of its recombinant adenovirus vector / 重庆医科大学学报

ABSTRACT

Objective:To construct the recombinant adenovirus vector carrying rat serum amyloid P component(SAP)as preparation for later use.Methods:Rat SAP was obtained by using RT-PCR amplification and then cloned into the shutter plasmid pAdTrace-TO4.Subsequently,this newly constructed plasmid pAdTrace-TO4-SAP was followed by homologous recombination with bone plasmid pAdEasy-1 in BJ5183.Recombinant adenovirus was obtained after being packaged in human embryonic kidney cells HEK293 by lipofectAmineTM2000 mediation.After amplification the recombined adenovirus,we determined its titer by end-point dilution assay.Results:In recombinant shuttle plasmid and recombinant adenovirus,a fragment of 700 bp detected by PCR was identical with that included in Genbank.The titer of recombinant adenovirus reached 2.8?108 pfu/ml.Conclusion:The recombinant adenovirus containing SAP gene was constructed successfully.This will provides a good basis for further study about the influence of SAP on lung fibrosis.