Quantitation of IFN-? mRNA in duck PBMC and its application
- VernacularTitle:外周血单个核细胞IFN-? mRNA的测定及应用
- Author:
Jianer LONG
;
Lina HUANG
;
Zhiqiang QIN
- Publication Type:Journal Article
- Keywords:
DuIFN-?;
Competitive RT-PCR;
DHBV;
DNA immunization
- From:
Chinese Journal of Infectious Diseases
2001;0(03):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective IFN-? is a pleiotropic cytokine with potent immunomodulatory effects and antiviral activity. To study the mechanism of IFN-? clearing duck hepatitis B virus (DHBV) in ducks, it is essential to establish a method to quantify expression of DuIFN-? in immune response. In the present study,a semi-quantitative competitive RT-PCR was developed to quantify expression of duck IFN-?(DuIFN-?) mRNA by PBMCs. Methods Based on ?-actin consensus sequence, fishing the ?-actin gene as house keeping gene from duck PBMC by RT-PCR. A competitive internal control was constructed and the competitive RT-PCR system could be used to quantify the transcription of DuIFN-? mRNA. Results After duck PBMCs were stimulated in vitro with PHA, the peak of DuIFN-? expression was at 24-36h. Then RT-PCR method was applied to detect DuIFN-? mRNA transcription by PBMCs from DHBV infected ducks immunized with DuIFN-? plasmid plus DNA vaccine or DNA vaccine alone. Results showed that expression of DuIFN-? in ducks co-immunized with DuIFN-? plasmid were higher than other groups immunized without DuIFN-? plasmid as adjuvant. Conclusions The results indicated that DuIFN-? gene could be a useful adjuvant to develop vaccines. The semi-quantitation of DuIFN-? mRNA by competitive RT-PCR provides the basis for future study of the mechanism of IFN-? in duck hepatitis B virus persistent infection.
