[Quantitative analysis of cell fusion caused by glycoprotein of paramyxoviruses with reporter gene method].
OBJECTIVE
To establish a kind of quantitative assay method for analyzing the reporter gene assay for fusion was established.


METHODS
Two populations of BHK21 cells were infected with vTF-7 recombinant and wild-type vaccinia viruses respectively and transfected with paramyxovirus F and HN cDNA and plasmid pG1NT7 beta-gal respectively. After 16 h, the two cell populations were removed from the wells by trypsinization. Equal numbers of the two populations were mixed in wells of a 96-well flat-bottom plate after washing and pelleting. After 15 hours' incubation at 37 degrees C, the cells were lysed with Nonidet P-40 and A values were read at 570 nm with an ELISA reader supported by SOFTMAX software.


RESULTS
Reporter gene method had a close relationship with cell count method, r = 0.9890(P < 0.01). Positive and negative coincidence rates were 100%. The best main reaction conditions are as follows: substrate concentration, 16 mmol/L; color reaction time, 20-25 min; amount of DNA for transfection, 1 microgram; cell numbers for each population, 1 x 10(5).


CONCLUSIONS
Reporter gene method is a kind of very sensitive, specific and repeatable quantitative analysis for cell fusion and can be used in the studies on membrane fusion caused by any viruses.
Wang
Z
Z
Department of Virology, Shandong Medical University, Jinan, 250012, China.
Yu
X
X
D002459:Cell Fusion; D017930:Genes, Reporter; D018098:Paramyxovirinae / Q000235:genetics* / Q000502:physiology; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000502:physiology
D014759:Viral Envelope Proteins