Adomas et al. BMC Genomics 2010, 11:477 http://www.biomedcentral.com/1471-2164/11/477 Page 10 of 16 Table 5 Validation of microarray and RNA sequencing of yeast grown in nitrogen poor (MM) and nitrogen rich (SC) conditions using Real Time RT PCR Microarray RNA sequencing RT PCR MTP oligo(dT) MTP oligo(dT) MTP Oligo(dT) Ratio MM/SC p value Ratio MM/SC p value Ratio MM/SC p value Ratio MM/SC p value Ratio MM/SC Ratio MM/SC Gene ID YDL244W YGR225W YGR249W YMR025W YNR072W YFR034C YKR053C YLR126C YMR003W YOR252W 1.98 1.72 1.66 1.51 -1.24 -1.42 -5.37 1.54 1.25 1.29 0.00 0.00 0.01 0.00 0.05 0.02 0.00 0.00 0.01 0.01 -1.25 -1.88 1.36 1.11 1.03 0.10 0.14 0.11 0.26 0.46 3.16 7.65 5.80 3.96 -6.32 -9.08 -2.16 5.54 7.91 6.33 0.11 0.00 0.00 0.15 0.00 0.00 0.00 0.05 0.01 0.00 -6.41 1.15 1.40 2.65 1.54 0.08 0.51 0.42 0.08 0.10 28.94 4.08 4.79 2.10 -2.34 1.30 -2.10 1.58 1.03 1.99 39.62 3.13 8.20 1.57 -1.36 1.24 -2.05 1.95 1.95 1.54 expression differences, whereas oligo-dT primers were not. Some genes feature multiple sites for priming by MTPs, leading to greater power for the detection of dif- ferential expression for those genes. However, the effect of multiple priming is modest, mostly increasing the chance of a gene being well measured. The equivalent coefficients of variation of our transcriptomic tag sequen- cing runs indicate that the complexity of the mRNA pool with either priming strategy is approximately the same. The advantage of adding MTPs obtains because the reverse transcription biases for each method of priming are different. In combination, they exhibit greater cover- age than either would individually. Based on our high- throughput transcript sequencing data, MTPs might yield slightly better coverage of expressed sequences than oligo-dT. In terms of false negatives in the high-through- put transcript sequencing, MTP priming exhibited a lower false-negative rate (248 not detected by MTP mix Figure 5 Genes significantly differentially expressed (P ≤ 0.05) by N. crassa protoperithecia (light red/blue) and mycelium (dark red/ blue) coding for enzymes involved in fatty acid metabolism; identified by microarray using multi-targeted primers (red) or oligo(dT) primers (blue). Error bars represent 95% credible intervals. NS - statistically insignificant difference (P > 0.05); * or lack of a symbol - significant difference (P ≤ 0.05); † - gene not well measured.