Methods of Reducing Repeat-Induced Silencing of Transgene Expression and Improved Fluorescent Biosensors

An isolated nucleic acid which encodes a ligand binding fluorescent indicator, the indicator comprising:

• a ligand binding protein moiety;
• a donor fluorophore moiety fused to the ligand binding protein moiety;
• and
• an acceptor fluorophore moiety fused to the ligand binding protein moiety;
• wherein fluorescence resonance energy transfer (FRET) between the donor moiety and the acceptor moiety is altered when the donor moiety is excited and said ligand binds to the ligand binding protein moiety, and
• wherein the nucleic acid sequence encoding at least one of either said donor fluorophore moiety or said acceptor fluorophore moiety has been genetically altered to reduce the level of nucleic acid sequence identity between the nucleic acid encoding the donor fluorophore moiety and the nucleic acid encoding the acceptor fluorophore moiety.