Direct calibration of microarray probes Making a DNA-meter Alex Pozhitkov, Peter A. Noble, Diethard Tautz, Ploen Workshop, 2011.

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Presentation transcript:

Direct calibration of microarray probes Making a DNA-meter Alex Pozhitkov, Peter A. Noble, Diethard Tautz, Ploen Workshop, 2011.

Conundrum Signal intensity vs. GC Signal intensity vs. G 0 Sense vs. antisense (same genomic loci) R 2 =0.37 Confirms: Pozhitkov, et al (2006). Tests of rRNA hybridization … cannot be predicted. NAR 34, e66-e6

Melting study Overhangs Buffer composition Tm Sense vs. Tm antisense Tm vs. Signal Intensity on the array

Melting Temperatures Tm Sense ( 0 C) Tm Antisense ( 0 C)

Microarray is an Instrument Instruments require calibration

Good, Bad and Noise Individual replicasAverages Concentration Signal Intensity

Freundlich isotherm! y=ax b Langmuir isotherm is not the only possible isotherm: Pozhitkov et al. (2010) Beyond Affymetrix arrays: … NAR 38, e28.

DNA and RNA y=ax b +c

Measurements and Errors Single probe replicateAverage over 10 replicates

CNV detection CNV expected: 1

Sample prep variability loci $

Acknowledgements Max Planck Society Alabama State University