A mobile gas chromatograph-mass spectrometer for measurement of metabolites and volatile substances in biological systems

Lead Research Organisation: University of Sheffield
Department Name: Animal and Plant Sciences

Abstract

To improve our ability to develop a sustainable agriculture that uses lower inputs and can respond to climate change requires experimentation in the field with agricultural crops. Such experiment require measurements of compounds such as metabolites, or compounds which are characteristic of organisms that live in close proximity to crops, and which provide important components for sustainable agriculture. Unfortunately many of these compounds are not very stable. Currently much expense and effort is required to transport samples to the laboratory for analysis, and it is not clear that the analyses that result accurately reflect the dynamics of plants in their field environment. . Field experiments need large amounts of replication because the field is not a very uniform environment. Mass spectrometry is a very powerful analytical tool to identify and measure these compounds but the machines tend to be very large. Thus until recently such field experiments have required expensive procedures to transport material back to the laboratory. However during the last 18 months a new mass spectrometer has become available. It is the size of a large brief case and can be taken to the field for the measurements required. The aim of this proposal is to purchase this machine so that the machine can be taken to the sample in the field, rather than the sample taken to the machine in the laboratory. This will improve both the accuracy and the efficiency of field experiments, while at the same time reducing costs associated with current sampling practice.
This new equipment will be maintained by the biOMICS Mass Spectrometry Facility in the Faculty of Science at the University of Sheffield.

Technical Summary

The aim of this application is to equip the biOMICS Mass Spectrometry Facility in the Faculty of Science at the University with a Torion field-portable gas chromatograph-mass spectrometer (GC-MS). The equipment combines solid-phase microextraction, purge-and-trap, thermal desorption, and heated headspace sampling techniques which provide a fast response for in-field analysis of organic compounds in a wide variety of samples including waters, plants, and soil. Two specific applications in current projects are the analysis of lipids which provide markers that allow the characterisation of microbial populations and the analysis of shoot exudates in different genotypes of maize. This purchase will allow the expertise in the Facility to be used directly in the field in numerous projects in which it is involved. The aim is to take the mass spectrometer to the sample rather than the sample to the mass spectrometer, thus improving the quality of analyses by undertaking both targeted and untargeted analyses in situ, with acquisition of quantitative and qualitative data from crops and plants that more accurately reflects their composition and dynamics at the time of sampling, while at the same time avoiding the costs of under-, over- or mis-sampling associated with the transport of samples back to a centralised Facility.

Planned Impact

This proposal is to enhance the capabilities of the Biomics Mass Spectrometry Facility in the Faculty of Science at the University of Sheffield by equipping it with the ability to make sensitive measurements associated with sustainable crop and food security programmes, using mass spectrometry out in the field.
The immediate benefit that will be gained is to extend the expertise of analytical biochemistry already present in the Facility to colleagues within the department working on projects involving sustainable agriculture as well as to associated colleagues at other Institutions. Research assistants and technicians on relevant projects will be trained in how to make measurements in the field and how to process and understand the data produced. A mobile GC-MS is designed to be used by the non-specialist once set up by a member of the Facility. However the metabolic profiling data acquired by the instrument requires complex statistical analysis and training will be provided to show how to analyse potentially very large data sets appropriately. The Facility also provides training in more fundamental aspects of mass spectrometry and analytical biochemistry which will guide colleagues in how to use the machine appropriately. Since the Facility supports a wide range of projects, some of which are in other Universities such as Leeds, Reading and York, the purchase of a mobile GC-MS will benefit these groups as above but also expose their colleagues to the potential of the new technology. The projects that the Facility supports involve making agriculture more sustainable, improving the quality of food and human health by increasing efficiency and sustainability of crop production. Thus in the longer term there will be significant benefits to society by improving quality of life.

Publications

10 25 50
 
Description This award allowed for the development of novel biological markers of plant stress
Exploitation Route We plan to publish our new methods and have received internal pump priming funds to apply those methods in a novel context with an industry partner
Sectors Agriculture

Food and Drink

Environment

 
Description We have received internal pump priming funds to apply those methods in a novel context with an industry partner (Severn Trent) to used the portable GCMS to test for damaging agricultural runoff entering the drinking water supply
First Year Of Impact 2019
Sector Agriculture, Food and Drink,Environment
Impact Types Societal

Economic

Policy & public services

 
Title Data for Efficient overexpression and purification of SARS-CoV-2 Nucleocapsid proteins in Escherichia coli 
Description The datasets provided relate to identification of minor contaminants found in recombinant native and 6-his-tagged SARS-CoV-2 Nucelocapsid proteins.  Abstract of publication: The fundamental biology of Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (Ncap), its use in diagnostic assays and its potential application as a vaccine component have received considerable attention since the outbreak of the Covid19 pandemic in late 2019. Here we report the scalable expression and purification of soluble, immunologically active, SARS-CoV-2 Ncap in Escherichia coli. Codon-optimised synthetic genes encoding the original Ncap sequence and four common variants with an N-terminal 6His affinity tag (sequence MHHHHHHG) were cloned into an inducible expression vector carrying a regulated bacteriophage T5 synthetic promoter controlled by lac operator binding sites. The constructs were used to express Ncap proteins and protocols developed which allow efficient production of purified Ncap with yields of over 200 mg per litre of culture media. These proteins were deployed in ELISA assays to allow comparison of their responses to human sera. Our results suggest that there was no detectable difference between the 6His-tagged and untagged original Ncap proteins but there may be a slight loss of sensitivity of sera to other Ncap isolates. 
Type Of Material Database/Collection of data 
Year Produced 2024 
Provided To Others? Yes  
URL https://orda.shef.ac.uk/articles/dataset/Data_for_Efficient_overexpression_and_purification_of_SARS-...