ProNet™ Quality Control

With over a decade of research experience in Yeast Two-Hybrid technology, we have developed a system which substantially reduces false positive interactions.

The ProNet quality control process

• Low copy number plasmids

  With fewer plasmids in the cell, we can keep the expression of selectable markers low, thus reducing the isolation of spurious interactions resulting from leaky expression

• Multiple reporter genes with dissimilar promoters

  This strategy reduces the possibility of promoter or reporter gene specific artifacts that might engender isolation of false positives
  

• Re-isolation of plasmids and co-transformation

  By removing both bait & prey plasmids from selected yeast interaction clones, transforming them into E. coli, cloning the plasmids yet again and co-transforming them both into yeast a 2nd time, we confirm the authenticity of a specific bait/prey interaction

• False positive test with heterologous protein baits

  Newly identified false positive, prey proteins that non-specifically associate with a wide variety bait proteins test are eliminated by our False Positive specificity assay. Here, we test the prey protein against a proprietary series of non-specific baits. Prey proteins from bonafide Bait/Prey interactions don’t associate with this panel of non-specific bait proteins, thus providing further assurance that the interaction is of a specific nature

• Statistical algorithm for removal of prevalent false positives

  During our years of experience with the Y2H process, we have developed a database encompassing those proteins which exhibit promiscuous binding behavior. These too, are eliminated by the ProNet quality control process.

   

  

   

  Only interactions surviving this rigorous set of criteria, are released as ProNet data.