#434 As a recent graduate of RMIT University you

As a recent graduate of RMIT University you - Biology

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As a recent graduate of RMIT University you have just been employed (3-year contract) by a world-renowned infectious diseases institute located in Victoria, Australia. You are required to investigate the effects of the novel small molecule anti-viral (DAD030981) to determine the global metabolic changes which take place in the tissue of lungs of monkeys infected by SARS- CoV-2 (Infected Group) vs the tissue of lungs of healthy monkeys (Control Group). You have been asked by your laboratory head to design and carry out a “metabolomics experiment.”

Q1. What experimental design would you propose to your lab head? He will first need to approve this and then discuss with you how you are going to prepare samples and analyse them before you begin.

What kind of metabolomics approach will you propose to your lab head and why? What considerations with respect to your experimental design should you propose?

Q2. Congratulations! Your lab head has approved your project and has now asked you to provide details as to how you are going to prepare samples for your metabolomics experiment.

Briefly describe your proposed sample preparation workflow ONLY and provide a justification for each step.

What kind of internal standard do you propose to use and why?
Q3. Your lab head has now asked you to analyse the polar metabolite extract on the laboratory’s GC-MS.

Briefly describe how it may be possible to analyse polar metabolites on the GC-MS. What additional sample should you run with you experiment and why?

Q4. Your lab head is quite pleased that you have acquired all data in such a short amount of time and is now eager for you to analyse the data.

Briefly explain what and why deconvolution is important in appropriately identifying metabolites.

How can your lab head determine which metabolites are up and/or down regulated? How can your lab head determine which metabolites are statistically significant?

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Dear student due to less time provided for solve we are advised to solve 1st Question when multiple Questions are uploaded (Chegg minimum answer policy). Your question is quite lengthy too. Hence i am solving the first part only. For the rest of the Question please upload it seperately.

Q 1) experimental design I would propose to my lab head would be -

Isolate the SARS COV-2 by aerosol or mucous fr mild clinical exposure . Here ,Factors that can affect the experiment have to be eliminated or identified so that they can be accounted for during data analysis. sample size, randomisation, and storage helps in producing reproducible and successful experiments that minimise erroneous variability.

Here ,There are two approaches in metabolomics:

1. Targeted approach

2. Untargeted approach

Targeted approach measures the set of metabolites to address specific biochemical question.

Untargeted approach, also called as global approach measures as many metabolites as possible from a range of biological samples without any bias.

• Workflow for both the approaches is as follows:
• Designing of the experiment.--> Sample Preparation --> Chromatography -->Mass Spectrometry -->Data acquisition --> Data processing --> Analysis and Imterpretation.

Sample Preparation:

It involves collection, storage, extraction and preparation of the sample for the individual measurement system

In most case, solid case extraction system is preferred in metabolomics studies.

Mass spectrometry (MS) :

It is an analytical technique used to measure small molecules.

The analytes are ionised at an ion source before they can be detected in a coupled mass detector.

Data processing:

Data processing aims to extract biologically relevant information from the acquired data. It includes many steps that are similar for MS. The random and systematic noise both are eliminated.

Analysis and Interpretation:It is done for categorisation , regression and prediction of the properties of the sample.