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Fig. 4. Quantitative comparison of amplified cDNA with unamplified cDNA. (A) The
relative gene expression level of each target gene in amplified cDNA (gray
column) was compared with that in unamplified cDNA (white column). Data
represent the average of three experiments. Error bars indicate s.d. (B)
Scatter plot showing the relationship between relative gene expression values
obtained from amplified and unamplified cDNA. Data obtained from
Fig. 3B were plotted. (C)
Reproducibility of cDNA amplification. Expression values of the melanogenic
genes were determined by Q-PCR using four replicates of amplified cDNA. The
expression value of each gene was consistent among the four amplifications.
(D) Detection of transcripts at known copy number in cDNA amplification.
Control poly(A)-tailed RNAs of known concentrations were mixed in lysis buffer
containing 20 pg total RNA prepared from Melb-a cells, reverse transcribed
followed by cDNA amplification. Four different RNA spikes, corresponding
roughly to 40, 400, 4000 and 40,000 copies/sample, were spiked into four
different tubes containing an identical amount of total RNA. Relative
expression values were determined by Q-PCR using a primer pair specific for
each spike RNA.
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