کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1936406 | 1050690 | 2008 | 8 صفحه PDF | دانلود رایگان |
![عکس صفحه اول مقاله: An accurate quantitative method for screening effective siRNA probes targeting a Hepatitis B virus transcript in single living cells An accurate quantitative method for screening effective siRNA probes targeting a Hepatitis B virus transcript in single living cells](/preview/png/1936406.png)
A dual fluorescence reporter plasmid expressing EGFP and DsRed-Monomer from separate promoters was constructed for quantitative flow cytometry analysis. Cloning the hepatitis B virus (HBV) X gene into the 3′ UTR region of DsRed-Monomer allowed quantifying the efficacy of ten siRNAs designed according to the accessibility of HBx mRNA measured in vitro. Using EGFP as an internal control, a justified calculation of the changed mean fluorescence intensity of DsRed-Monomer in each transfected cell yielded highly consistent results, and revealed all 10 siRNAs achieved over 50% inhibition among which a super effective siRNA achieved 88% inhibition at a very low concentration (0.33 μg/ml). This provides a quantification method critical for therapeutic application of siRNA.
Journal: Biochemical and Biophysical Research Communications - Volume 367, Issue 4, 21 March 2008, Pages 866–873