TY - JOUR
T1 - Directed Evolution of Transcription Factor-Based Biosensors for Altered Effector Specificity
AU - Machado, Leopoldo Ferreira Marques
AU - Dixon, Neil
N1 - © 2022. Springer Science+Business Media, LLC, part of Springer Nature.
PY - 2022
Y1 - 2022
N2 - Transcription factor-based biosensors are important tools in Synthetic Biology for the sensing of industrially valuable molecules and clinically important metabolites, therefore presenting applications in the bioremediation, industrial biotechnology, and biomedical fields. The directed evolution of allosteric transcription factors (aTFs) with the aim of altering effector specificity has the potential for the development of new biosensors to detect natural and nonnatural molecules, expanding the scope of available aTF-based biosensors. In this chapter, we delineate a general method for the directed evolution of aTFs. The theory of library design is discussed, along with the detailed methodology for an improved transformation of combined libraries, and the experimental search space by counterselection using fluorescence-activated cell sorting (FACS) is presented.
AB - Transcription factor-based biosensors are important tools in Synthetic Biology for the sensing of industrially valuable molecules and clinically important metabolites, therefore presenting applications in the bioremediation, industrial biotechnology, and biomedical fields. The directed evolution of allosteric transcription factors (aTFs) with the aim of altering effector specificity has the potential for the development of new biosensors to detect natural and nonnatural molecules, expanding the scope of available aTF-based biosensors. In this chapter, we delineate a general method for the directed evolution of aTFs. The theory of library design is discussed, along with the detailed methodology for an improved transformation of combined libraries, and the experimental search space by counterselection using fluorescence-activated cell sorting (FACS) is presented.
KW - Biosensing Techniques/methods
KW - Biotechnology
KW - Flow Cytometry
KW - Gene Library
KW - Transcription Factors/genetics
U2 - 10.1007/978-1-0716-2152-3_12
DO - 10.1007/978-1-0716-2152-3_12
M3 - Article
C2 - 35727451
SN - 1064-3745
VL - 2461
SP - 175
EP - 193
JO - Methods in molecular biology (Clifton, N.J.)
JF - Methods in molecular biology (Clifton, N.J.)
ER -
