Dr Adil Khan is a postdoctoral researcher in the laboratory of Prof Ryan Lister at the University of Western Australia. Adil Khan completed his Master in Biotechnology (Research) at The Australian National University and subsequently completed his PhD in 2021 in Plant Synthetic Biology at The University of Western Australia. As a graduate student, Adil worked with Prof Ryan Lister to develop reversible CRISPRi circuits for reprogramming gene expression in plants. Adil is continuing as a postdoctoral researcher in the laboratory of Prof Ryan Lister to develop new tools for reprogramming plants
Monday session 5
CRISPRi-based circuits for genetic computation in plants
A. KHAN1, G. HERRING1, M. OLIVA1, E. FOURIE1, J. Y. ZHU1, B. JOHNSTON1, J. POTTER1, J. PFLUGER2, T. SWAIN2, C. PFLUGER2, J. LLOYD1, D. SECCO1, I. SMALL1, B. KIDD1,3, R. LISTER1,2
1Australian Research Council Centre of Excellence in Plant Energy Biology, School of Molecular Sciences, The University of Western Australia, WA, Australia 2Harry Perkins Institute of Medical Research, The University of Western Australia, WA, Australia 3CSIRO Synthetic Biology Future Science Platform, QLD, Australia
Synthetic gene circuits can enable new cellular behaviours by integrating multiple input signals into customisable genetic programs. However, gene circuit development in plants has been limited by a lack of orthogonal and modular parts required for their construction. Here, we present the first implementation of reversible CRISPRi-based gene circuits in plants. First, we created a toolkit of engineered repressible promoters of different strengths and used them as integrators for the construction of NOT and NOR gates in Arabidopsis thaliana. Next, we determined the optimal processing system to express sgRNAs from RNA Pol II promoters to introduce NOR gate programmability for interfacing with host regulatory sequences. Finally, we layered multiple NOR gates together to create OR, NIMPLY, and AND logic functions and tested NOR gate activity in Physcomitrium patens to demonstrate cross-species utility. Our CRISPRi circuits are orthogonal, compact, reversible, programmable, and modular, providing a new platform for sophisticated spatio-temporal control of gene expression in plants.