After DNase1 treatment (Thermo Scientific), cDNA was synthesized
using the RevertAid Reverse Transcriptase (Thermo Scientific). Quantitative
real-time PCR was performed using Power SYBR Green (Applied Biosystems) in a 5 μl
reaction using the standard program of a ViiA™ 7 instrument (Applied Biosystems).
Data was extracted using ViiA™ 7 Software v1.1. Primer amplification efficiency
was calculated using LinRegPCR [65 (link)].
Primers used are listed in Additional file10 : Table S5. Results for the primer recognizing the luciferase
RNA spike were used to normalize real time qRT-PCR data and the values obtained
from gradient fraction samples were reported to the area under the corresponding
gradient A254 curve to adjust for different RNA contents of the
samples before extraction.
using the RevertAid Reverse Transcriptase (Thermo Scientific). Quantitative
real-time PCR was performed using Power SYBR Green (Applied Biosystems) in a 5 μl
reaction using the standard program of a ViiA™ 7 instrument (Applied Biosystems).
Data was extracted using ViiA™ 7 Software v1.1. Primer amplification efficiency
was calculated using LinRegPCR [65 (link)].
Primers used are listed in Additional file
RNA spike were used to normalize real time qRT-PCR data and the values obtained
from gradient fraction samples were reported to the area under the corresponding
gradient A254 curve to adjust for different RNA contents of the
samples before extraction.
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