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Dr Patrick Hardinge
Research Fellow
School of Biosciences
- HardingeP@cardiff.ac.uk
- Sir Martin Evans Building, Room Room W/3.12, Museum Avenue, Cardiff, CF10 3AX
Overview
My main interest is the detection of trace amounts of DNA in biological samples. This stems from the years I spent working as a Forensic Scientist and following the basic principle of forensic science that ‘every contact leaves a trace’. I specialise in the application of isothermal DNA amplification, predominantly loop-mediated amplification (LAMP), to increase the trace target DNA to a level that is detectable. I work primarily with bioluminescent detection and have a long established collaboration with ERBA MDX in Ely, Cambridgeshire and I use a range of imaging techniques to visualise and record the results. I have also developed and researched alternative fluorescent detection methods for LAMP DNA amplification which have been published recently.
My current research project allows me to explore new areas with an exciting collaboration within Cardiff University with the Schools of Pharmacy and Engineering, to use microfluidics to produce droplets and artificial cells containing a bioluminescent DNA amplification reaction. The stability of the multiple uniform droplets opens the possibility of digital LAMP quantification and the bioluminescent reaction allows us to study the artificial cells.
Also in the project I have developed sequencing and bioinformatics skills in the de novo assembly of a fluke that is of increasing veterinary interest in the UK. The main aim of this work is to develop a specific, sensitive test to detect this infection and combine this research with microfluidic droplet generation and amplification detection.
My research is supported by the Welsh Government’s Sêr Cymru II programme which is part funded by Cardiff University and the Welsh European Funding Office (WEFO) under the European Regional Development Fund (ERDF).
Publication
2023
- Hardinge, P. 2023. Molecular beacons - loop-mediated amplification (MB-LAMP). In: Shavrukov, Y. ed. Plant Genotyping: Methods and Protocols., Vol. 2638. Methods in Molecular Biology New York, US: Springer, pp. 289-299., (10.1007/978-1-0716-3024-2_20)
2022
- Hardinge, P. 2022. Optimized loop-mediated isothermal amplification (lamp) allows single copy detection using bioluminescent assay in real time (bart). In: Bioluminescence: Methods and Protocols., Vol. 2524. Springer, pp. 107-117., (10.1007/978-1-0716-2453-1_8)
- Nan, X. et al. 2022. VarLOCK - sequencing independent, rapid detection of SARS-CoV-2 variants of concern for point-of-care testing, qPCR pipelines and national wastewater surveillance. [Online]. medRxix: (10.1101/2022.01.06.21268555) Available at: https://www.medrxiv.org/content/10.1101/2022.01.06.21268555v1
2021
- Hardinge, P., Baxani, D. K., McCloy, T., Murray, J. A. H. and Castell, O. K. 2021. Author Correction: Bioluminescent detection of isothermal DNA amplification in microfluidic generated droplets and artificial cells. Scientific Reports 11(1), article number: 10748. (10.1038/s41598-021-90099-5)
2020
- Hardinge, P., Baxani, D. K., McCloy, T., Murray, J. A. H. and Castell, O. K. 2020. Bioluminescent detection of isothermal DNA amplification in microfluidic generated droplets and artificial cells. Scientific Reports 10(1), article number: 21886. (10.1038/s41598-020-78996-7)
- Hardinge, P. and Murray, J. A. H. 2020. Full dynamic range quantification using loop-mediated amplification (LAMP) by combining analysis of amplification timing and variance between replicates at low copy number. Scientific Reports 10, article number: 916. (10.1038/s41598-020-57473-1)
2019
- Hardinge, P. and Murray, J. A. H. 2019. Lack of specificity associated with using molecular beacons in loop mediated amplification assays. BMC Biotechnology 19, article number: 55. (10.1186/s12896-019-0549-z)
- Hardinge, P. and Murray, J. A. H. 2019. Reduced false positives and improved reporting of loop-mediated isothermal amplification using quenched fluorescent primers. Scientific Reports 9, article number: 7400. (10.1038/s41598-019-43817-z)
2018
- Hardinge, P., Kiddle, G., Tisi, L. and Murray, J. A. H. 2018. Optimised LAMP allows single copy detection of 35Sp and NOSt in transgenic maize using Bioluminescent Assay in Real Time (BART). Scientific Reports 8, article number: 17590. (10.1038/s41598-018-36207-4)
2014
- Hardinge, P. 2014. Low copy number quantification of DNA utilising Loop-mediated Amplification (LAMP) with Bioluminescent Assay in Real-Time (BART) reporter. PhD Thesis, Cardiff University.
2013
- Hardinge, P., Allard, J., Wain, A. and Watson, S. 2013. Optimisation of choline testing using Florence Iodine reagent, including comparative sensitivity and specificity with PSA and AP tests. Science & Justice 53(1), pp. 34-40. (10.1016/j.scijus.2012.03.004)
2012
- Kiddle, G. et al. 2012. GMO detection using a bioluminescent real time reporter (BART) of loop mediated isothermal amplification (LAMP) suitable for field use. BMC Biotechnology 12, article number: 15. (10.1186/1472-6750-12-15)
Articles
- Hardinge, P., Baxani, D. K., McCloy, T., Murray, J. A. H. and Castell, O. K. 2021. Author Correction: Bioluminescent detection of isothermal DNA amplification in microfluidic generated droplets and artificial cells. Scientific Reports 11(1), article number: 10748. (10.1038/s41598-021-90099-5)
- Hardinge, P., Baxani, D. K., McCloy, T., Murray, J. A. H. and Castell, O. K. 2020. Bioluminescent detection of isothermal DNA amplification in microfluidic generated droplets and artificial cells. Scientific Reports 10(1), article number: 21886. (10.1038/s41598-020-78996-7)
- Hardinge, P. and Murray, J. A. H. 2020. Full dynamic range quantification using loop-mediated amplification (LAMP) by combining analysis of amplification timing and variance between replicates at low copy number. Scientific Reports 10, article number: 916. (10.1038/s41598-020-57473-1)
- Hardinge, P. and Murray, J. A. H. 2019. Lack of specificity associated with using molecular beacons in loop mediated amplification assays. BMC Biotechnology 19, article number: 55. (10.1186/s12896-019-0549-z)
- Hardinge, P. and Murray, J. A. H. 2019. Reduced false positives and improved reporting of loop-mediated isothermal amplification using quenched fluorescent primers. Scientific Reports 9, article number: 7400. (10.1038/s41598-019-43817-z)
- Hardinge, P., Kiddle, G., Tisi, L. and Murray, J. A. H. 2018. Optimised LAMP allows single copy detection of 35Sp and NOSt in transgenic maize using Bioluminescent Assay in Real Time (BART). Scientific Reports 8, article number: 17590. (10.1038/s41598-018-36207-4)
- Hardinge, P., Allard, J., Wain, A. and Watson, S. 2013. Optimisation of choline testing using Florence Iodine reagent, including comparative sensitivity and specificity with PSA and AP tests. Science & Justice 53(1), pp. 34-40. (10.1016/j.scijus.2012.03.004)
- Kiddle, G. et al. 2012. GMO detection using a bioluminescent real time reporter (BART) of loop mediated isothermal amplification (LAMP) suitable for field use. BMC Biotechnology 12, article number: 15. (10.1186/1472-6750-12-15)
Book sections
- Hardinge, P. 2023. Molecular beacons - loop-mediated amplification (MB-LAMP). In: Shavrukov, Y. ed. Plant Genotyping: Methods and Protocols., Vol. 2638. Methods in Molecular Biology New York, US: Springer, pp. 289-299., (10.1007/978-1-0716-3024-2_20)
- Hardinge, P. 2022. Optimized loop-mediated isothermal amplification (lamp) allows single copy detection using bioluminescent assay in real time (bart). In: Bioluminescence: Methods and Protocols., Vol. 2524. Springer, pp. 107-117., (10.1007/978-1-0716-2453-1_8)
Thesis
- Hardinge, P. 2014. Low copy number quantification of DNA utilising Loop-mediated Amplification (LAMP) with Bioluminescent Assay in Real-Time (BART) reporter. PhD Thesis, Cardiff University.
Websites
- Nan, X. et al. 2022. VarLOCK - sequencing independent, rapid detection of SARS-CoV-2 variants of concern for point-of-care testing, qPCR pipelines and national wastewater surveillance. [Online]. medRxix: (10.1101/2022.01.06.21268555) Available at: https://www.medrxiv.org/content/10.1101/2022.01.06.21268555v1
Biography
Before my PhD at Cardiff University I spent over a decade working as a Forensic Scientist in research and development and on cases for the criminal justice system at the Forensic Science Service. During my PhD and subsequent post-doctoral research at Cardiff University I focused on DNA quantification with fluorescent and bioluminescent detection.
Sêr Cymru II Fellowship project:
Development, evaluation and validation of a microfluidic device for pathogen detection using isothermal DNA amplification and bioluminescent detection. (Molecular Biology)
The project combines isothermal amplification of target DNA molecules with the bioluminescent assay in real time (BART), in collaboration with ERBA MDX, using biochemistry derived from fireflies. My PhD and post-doctoral research led me to the detection and quantification of very low numbers of target DNA molecules and fuelled an interest in developing a full dynamic range quantification strategy capable of determining concentrations from very low to very high with accuracy. The project enables me to collaborate with experts from the Schools of Pharmacy and Engineering at Cardiff University in the exciting field of microfluidics. The incorporation of bioluminescent reagents into multiple numbers of tiny uniform droplets provides me with valuable quantification data from light detection. The development of a simple device that brings together the benefits of isothermal DNA amplification, BART bioluminescent detection, microfluidic droplet generation and encapsulation with light detection and analysis, is the main focus of my research. I am also interested in the development of thermostable luciferase enzymes from fireflies for the BART detection and in the development of the target DNA detection towards a number of parasites of veterinary interest. More recently I have developed an interest in genomics, Oxford Nanopore Technology MinION sequencing and bioinformatics for de novo genome assembly.