Gene editing

The Genome Innovation Hub continually develops a number of protocols and procedures while undertaking our collaborative projects. Below are a selection of detailed developments useful to researchers undertaking Gene editing procedures in the laboratory.

Developments

Cas9 protein expression, purification and validation

CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR associated) technologies enable scientists to accurately modify the genome of cells and organisms. CRISPR-Cas9 and CRISPR-Cas12a enzymes are the most widely studied of the RNA-guided endonucleases used for genome editing. To help reduce costs of our genome editing research projects, we have established an "in-house" protocol for purifiying and functionally testing a range of Cas9/Cas12a endonuclease which are attached to various affinity tags.

Genome editing of mammalian cells using RNP

Currently in development at GIH is a basic workflow of genome editing in mammalian cells including guide RNA design, transfection using electroporation method to deliver RNP to the cell and editing analysis using a T7E1 assay. Commercial source of Cas9 and synthetic guide RNA was used to eliminate the workload of sub-cloning or in vitro transcription of guide RNA and achieve a high efficiency.

Featured projects	Duration
Optimised bioinformatics and validation pipeline for genome-wide CRISPR screening data Genome Innovation Hub Collaborative Project (UQ infrastructure)	2021
Expanding the scope of multimodal single cell sequencing Genome Innovation Hub Collaborative Project (UQ infrastructure)	2021
HTS single cell CRISPR technology for experimental validation of population genetics and functional genomics discovery Genome Innovation Hub Collaborative Project (UQ infrastructure)	2021
OmniCRISPR: a simultaneous and combinatorial approach to the generation of multiple precise genomic alterations Genome Innovation Hub Collaborative Project (UQ infrastructure)	2020
Simultaneous identification of RNA-chromatin interactions and transcriptomes in single cells Genome Innovation Hub Collaborative Project (UQ infrastructure)	2020
Genome-wide CRISPR screening for modifiers of diverse cellular phenotypes Genome Innovation Hub Collaborative Project (UQ infrastructure)	2019
Using transposon-sequencing to probe whole cell protein-protein interactions Genome Innovation Hub Collaborative Project (UQ infrastructure)	2019
Precise genome editing Genome Innovation Hub Collaborative Project (UQ infrastructure)	2019
Dual gRNA screen Genome Innovation Hub Collaborative Project (UQ infrastructure)	2019

Publications

Therapeutic Inhibition of Acid-Sensing Ion Channel 1a Recovers Heart Function After Ischemia–Reperfusion Injury

Meredith A. Redd, PhD; Sarah E. Scheuer, MD; Natalie J. Saez, PhD; Yusuke Yoshikawa, MD, PhD; Han Sheng Chiu, MS; Ling Gao, PhD; Mark Hicks, PhD; Jeanette E. Villanueva, PhD; Yashutosh Joshi, MD; Chun Yuen Chow, PhD; Gabriel Cuellar-Partida, PhD; Jason N. Peart, PhD; Louise E. See Hoe, PhD; Xiaoli Chen, PhD; Yuliangzi Sun, MS; Jacky Y. Suen, PhD; Robert J. Hatch, PhD; Ben Rollo, PhD; Di Xia, PhD; Mubarak A.H. Alzubaidi, PhD; Snezana Maljevic, PhD; Gregory A. Quaife-Ryan, PhD; James E. Hudson, PhD; Enzo R. Porrello, PhD; Melanie Y. White, PhD; Stuart J. Cordwell, PhD; John F. Fraser, MD, PhD; Steven Petrou, PhD; Melissa E. Reichelt, PhD; Walter G. Thomas, PhD; Glenn F. King, PhD; Peter S. Macdonald, MD, PhD; Nathan J. Palpant, PhD

doi: https://www.ahajournals.org/doi/10.1161/CIR.0000000000001020

Cavin4 interacts with Bin1 to promote T-tubule formation and stability in developing skeletal muscle.

Harriet P. Lo, Ye-Wheen Lim, Zherui Xiong, Nick Martel, Charles Ferguson, Nicholas Ariotti, Jean Giacomotto, James Rae, Matthias Floetenmeyer, Shayli Varasteh Moradi, Ya Gao, Vikas A. Tillu, Di Xia, Huang Wang, Samira Rahnama, Susan J. Nixon, Michele Bastiani, Ryan D. Day, Kelly A. Smith, Nathan J. Palpant, Wayne A. Johnston, Kirill Alexandrov, Brett M. Collins, Thomas E. Hall, Robert G. Parton; Cavin4 interacts with Bin1 to promote T-tubule formation and stability in developing skeletal muscle. J Cell Biol 6 December 2021; 220 (12): e201905065.

doi: https://doi.org/10.1083/jcb.201905065

Dr Seth Cheetham

Honorary Fellow

Mater Research Institute-UQ, Faculty of Medicine
Joanna Crawford

Senior Research Assistant

Institute for Molecular Bioscience
Professor Geoff Faulkner

Professor in Neuroscience

Queensland Brain Institute
Affiliate Professor

Mater Research Institute-UQ, Faculty of Medicine
Dr Emily Goodall

Research Officer

Institute for Molecular Bioscience
Professor Ian Henderson

Director

Institute for Molecular Bioscience
Affiliate Professor

UQ Centre for Clinical Research, Faculty of Medicine
Affiliate Professor

School of Chemistry and Molecular Biosciences, Faculty of Science
Dr Jun Ma

Research Assistant

Genome Innovation Hub
Dr Dmitry Ovchinnikov

Senior Research Fellow

The Florey Institute of Neuroscience and Mental Health Melbourne, Brain Centre/Kenneth Myer Building, The University of Melbourne
Dr Nathan Palpant

Senior Research Fellow - GL

Institute for Molecular Bioscience
Dr Rebecca San Gil

FightMND Research Fellow

Queensland Brain Institute
Dr Adam Walker

Ross Maclean Fellow

Queensland Brain Institute
Isabel Warner

PhD candidate

Institute for Molecular Bioscience
Dr Qiongyi Zhao

Research Fellow

Queensland Brain Institute

Research

Gene editing

Developments

Cas9 protein expression, purification and validation

Genome editing of mammalian cells using RNP

Publications