Team:Sydney Australia/Parts

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Parts Overview

Introduction

This year we have made 11 parts to contribute to the iGEM parts registry and improved the characterisation of one existing University of Sydney part. The functions of our parts range from fluorescence to psychotropic drug formation and we’re thrilled by how much we’ve been able to accomplish in such a short amount of time.

Our Favourite Parts!

We know they say you’re not supposed to have favourites… but these are parts not children so we have favourites.


Name Part Number Function Why we love it

"I loved this part like a son. It was a father to me."
VVD36-C73A-CH4-M130I BBa_K3140010 An Light-Oxygen-Voltage domain (LOV) fluorescent protein 456 bp long generated from a CH4-codon optimised vvd gene (BBa_K3140009) with the conserved cysteine residue to alanine mutation in the LOV domain (C73A) and by introducing a point mutation from methione to isoleucine (M130I). This results in a fluorescent protein roughly 7 times the brightness compared to the wild-type (BBa_K3140005).We love how with just a couple of amino acid changes you can get a super bright fluorescent protein from a blue light sensor protein - doesn’t that just showcase the power of Synthetic biology and the awesome power of error prone PCR.

Benj: “I have a complicated relationship with this part…”

PsiD - Tryptophan decarboxylase from Psilocybe cubensis BBa_K3140000 An L-tryptophan decarboxylase. PsiD can convert tryptophan into tryptamine. We have been able to characterise the sequence, protein size, and confirm its activity by performing an enzyme assay and sending the product off for LCMS.This part almost never came to be. After many weeks of purification, PCR, and transformations we discovered that our main transformant had a frame-shifting deletion! We thought all hope was lost, but luckily we are plate hoarders and could test more of our initial transformations for those deletions and got great sequencing data that lead us to the part we have today. No one ever said that this would be easy!
PsiK - 4-hydroxytryptamine kinase from Psilocybe cubensis BBa_K3140002 A 4-hydroxytryptamine kinase. PsiH converts 4-hydroxytryptamine to norbaeocystin. We have been able to characterise this enzyme’s DNA sequence, protein size, and confirm its activity with LCMS. This enzyme officially makes our e.coli magical, as it produces the first psychedelic compound in our biosynthetic pathway. We were absolutely thrilled to receive this LCMS data so it had to become one of our favourite parts.

Our Second Favourites...

Name Part Number Function
PsiH - Cytochrome P450 monooxygenase from Psilocybe cubensis BBa_K3140001 A mono-oxygenase that can theoretically convert tryptamine to 4-hydroxytryptamine. We have sequencing and protein size data for this construct. We are waiting on conclusive LCMS data (come to our presentation!).
PsiM - Norbaeocystin methyltransferase from Psilocybe cubensis BBa_K3140003 A methyl-transferase which can convert norbaeocystin to baeocystin and theoretically convert baeocystin to psilocybin. We have sequencing, protein size data, and putative enzyme activity data for this enzyme.
PsiH (codon optimised) - Cytochrome P450 monooxygenase from Psilocybe cubensis BBa_K3140004 This part is an IDT-codon optimised version of our WT-PsiH part (listed above).
VVD36-C73A - Truncated VIVID fluoroprotein derived from Neurospora crassa BBa_K3140005 A blue fluorescent protein generated from mutating an existing blue light sensor part from the registry (Part:BBa_K1616014). We have sequencing and fluorescence data for this protein.
VVD36-C73A-CH1 (rank order harmonised) BBa_K3140006 A blue fluorescent protein made as a codon harmonised variant of VVD_wt using the rank order method. We have sequencing and fluorescence data for this protein.
VVD36-C73A-CH2 (nearest frequency harmonised) BBa_K3140007 A blue fluorescent protein generated as an absolute frequency codon harmonised variant of VVD_wt. We have sequencing and fluorescence data for this protein.
VVD36-C73A-CH3 (relative proportion harmonisation) BBa_K3140008 A blue fluorescent protein generated as a codon harmonised version of VVD_wt using the relative frequency method. We have sequencing and fluorescence data for this protein.
VVD36-C73A-CH4 (codon optimised) BBa_K3140009 A blue fluorescent protein generated as an IDT-codon optimised variant of VVD_wt. It is also the basis from which VVD_130 was made. We have sequencing and fluorescence data for this protein.