text: In any study, some of the protein domains, those under investigation, will be varied according to the goals of the study whereas other domains, those that are not themselves being investigated, will be kept constant. For example, in a two-hybrid study to select DNA-binding domains, the DNA-binding domain, BD, will be varied while the two interacting proteins, the bait and prey, must be kept constant to maintain a strong binding between the BD and AD. There are a number of domains from which to choose the BD, bait and prey and AD, if these are to remain constant. In protein–protein interaction investigations, the BD may be chosen from any of many strong DNA-binding domains such as Zif268. A frequent choice of bait and prey domains are residues 263–352 of yeast Gal11P with a N342V mutation and residues 58–97 of yeast Gal4, respectively. These domains can be used in both yeast- and bacterial-based selection techniques and are known to bind together strongly.
  The AD chosen must be able to activate transcription of the reporter gene, using the cells own transcription machinery. Thus, the variety of ADs available for use in yeast-based techniques may not be suited to use in their bacterial-based analogues. The herpes simplex virus-derived AD, VP16 and yeast Gal4 AD have been used with success in yeast whilst a portion of the α-subunit of E. coli RNA polymerase has been utilised in E. coli'-based methods.
  Whilst powerfully activating domains may allow greater sensitivity towards weaker interactions, conversely, a weaker AD may provide greater stringency.