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Jiajian Liu and Gary D. Stormo Presented by Aliya Sadeque

Combining SELEX with quantitative assays to rapidly obtain accurate models of protein–DNA interactions. Jiajian Liu and Gary D. Stormo Presented by Aliya Sadeque. Protein-DNA interactions. Methods for measuring: Yeast 1 hybrid ChIP on chip DNA microarray

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Jiajian Liu and Gary D. Stormo Presented by Aliya Sadeque

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  1. Combining SELEX with quantitative assays to rapidly obtain accurate models of protein–DNA interactions Jiajian Liu and Gary D. Stormo Presented by Aliya Sadeque

  2. Protein-DNA interactions • Methods for measuring: • Yeast 1 hybrid • ChIP on chip • DNA microarray • Important distinction in terms of specificity • Enzymes vs. transcription factors Bioinformatics. 2000 Jan;16(1):16-23.

  3. TFBSTranscription Factor Binding Sites • Knowing the specificity of a TF in order to locate its binding sites within the genome • Databases • TRANSFAC, JASPAR • Represented as consensus sequences or weight matrices

  4. SELEXSystematic Evolution of Ligands By Exponential Enrichment http://rulai.cshl.edu/tools/ESE2/ESEmatrix.html

  5. QuMFRA Nucleic Acids Res. 2001 Jun 15;29(12):2471-8. PMID: 11410653

  6. Selling points • Provides a general method that can be used for any DNA-binding protein even if nothing is known about its specificity. • Can isolate a small set of specific binding sites from a very large pool of random sequences Nucleic Acids Res. 2001 Jun 15;29(12):2471-8.

  7. Zif268

  8. SELEX Procedure

  9. Estimating Affinities • Assuming an additive model • Frequency: • Weight

  10. Sequence Logo

  11. QuMFRA Procedure • 15 sequences • Cover the space of possible sequence http://www.answers.com/topic/gel-abpp-eg-png

  12. Uh oh, math.

  13. critiques • we used affinity chromatography to modify the conventional SELEX procedure to save time and labor. –didn’t explain how • as determined by sequencing only ∼20 selected products after each round. –why only 20? • Why no A at position 1 in QuMFRA? • Discussion brought up weakness in SELEX

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