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OUR PARTS

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Basic Parts

During this project, our team designed the following basic parts.

Plasmid backbones

PartDescription
BBa_K5061001Helper phage (HP) M13KO7-ΔgVI backbone
BBa_K5061101Helper phage (HP) M13KO7-ΔgIII-ΔgVI backbone
BBa_K5061002Accessory Plasmid (AP) and Test Plasmid (TP) backbone
BBa_K5061003Mutagenesis Plasmid (MP) & Expression Plasmid (EP) backbone

Coding sequences

PartDescription
BBa_K5061011M13 bacteriophage gIII G378S
BBa_K5061012M13 bacteriophage gVI
BBa_K5061013LuxAB fusion protein
BBa_K5061060XylS
BBa_K5061061XylS K38R L224Q
BBa_K5061062XylS W88C L224Q
BBa_K5061063XylS AI PA
BBa_K5061064XylS AI TPA
BBa_K5061065XylS PHAGEVO-AI v1 PA
BBa_K5061066XylS PHAGEVO-AI v1 TPA
BBa_K5061067XylS PHAGEVO-AI v2 PA
BBa_K5061068XylS PHAGEVO-AI v2 TPA
BBa_K5061069XylS PHAGEVO-AI v3 r3m8 PA
BBa_K5061070XylS PHAGEVO-AI v3 r5m8 PA
BBa_K5061071XylS PHAGEVO-AI v3 r5m10 PA
BBa_K5061072XylS PHAGEVO-AI v3 r5m11 PA
BBa_K5061073XylS PHAGEVO-AI v3 r6m1 TPA
BBa_K5061074XylS PHAGEVO-AI v3 r6m5 TPA
BBa_K5061075XylS PHAGEVO-AI v3 r7m1 PA

Ribosomal binding sites (RBS)

PartDescription
BBa_K5061021Synthetic RBS designed for M13 gI
BBa_K5061022Synthetic RBS designed for M13 gIII
BBa_K5061023Synthetic RBS designed for M13 gVI
BBa_K5061024Synthetic RBS designed for XylS

Golden Gate adapters

PartDescription
BBa_K5061005Golden Gate adapter for Helper phages (HP) M13KO7-ΔgVI and M13KO7-ΔgIII-ΔgVI Golden Gate cloning platforms
BBa_K5061006Golden Gate adapter for Accessory Plasmids (AP) Golden Gate cloning platforms for M13 gVI expression
BBa_K5061007Golden Gate adapter for Test Plasmid (TP) Golden Gate cloning platform
BBa_K5061008Golden Gate adapter for Expression Plasmid (EP) Golden Gate cloning platform

and one Promoter

PartDescription
BBa_K5061010PspA promoter

Composite Parts

During this project, our team designed the following composite parts.

PHAGEVO Golden Gate cloning platforms

PartDescription
BBa_K5061031Helper phage (HP) M13KO7-ΔgVI Golden Gate cloning platform
BBa_K5061131Helper phage (HP) M13KO7-ΔgIII-ΔgVI Golden Gate cloning platform
BBa_K5061032Accessory Plasmid (AP) Golden Gate cloning platform for M13 gVI expression
BBa_K5061132Accessory Plasmid (AP) Golden Gate cloning platform for M13 gIII and gVI expression
BBa_K5061033Expression Plasmid (EP) Golden Gate cloning platform
BBa_K5061034Test Plasmid (TP) Golden Gate cloning platform

PHAGEVO AP, TP and MP plasmids

PartDescription
BBa_K5061041Mutagenesis Plasmid (MP) expressing the evoCDA1-BE4max - T7RNAP fusion protein from the pBad promoter
BBa_K5061042Mutagenesis Plasmid (MP) expressing the ABE8.20-m - T7RNAP fusion protein from the pBad promoter
BBa_K5061043Mutagenesis Plasmid (MP) expressing the evoCDA1-BE4max - T7RNAP-CGG-R12-KIRV fusion protein from the pBad promoter
BBa_K5061044Mutagenesis Plasmid (MP) expressing the ABE8.20-m - T7RNAP-CGG-R12-KIRV fusion protein from the pBad promoter
BBa_K5061045Accessory Plasmid (AP) expressing the M13 gVI and LuxAB from the PspA promoter
BBa_K5061145Accessory Plasmid (AP) expressing the M13 gIII, M13 gVI and LuxAB from the PspA promoter
BBa_K5061046Test Plasmid (TP) expressing XylS AI TPA from the pTAC promoter

PHAGEVO HP, AP, EP and TP plasmids for XylS evolution and testing

Part Description
BBa_K5061051 Helper phage (HP) M13KO7-ΔgVI carrying XylS K38R L224Q
BBa_K5061151 Helper phage (HP) M13KO7-ΔgIII-ΔgVI carrying XylS K38R L224Q
BBa_K5061052 Helper phage (HP) M13KO7-ΔgVI carrying XylS K38R L224Q in the PHAGEVO mutational region
BBa_K5061152 Helper phage (HP) M13KO7-ΔgIII-ΔgVI carrying XylS K38R L224Q in the PHAGEVO mutational region
BBa_K5061053 Helper phage (HP) M13KO7-ΔgVI carrying XylS W88C L224Q
BBa_K5061153 Helper phage (HP) M13KO7-ΔgIII-ΔgVI carrying XylS W88C L224Q
BBa_K5061054 Helper phage (HP) M13KO7-ΔgVI carrying XylS W88C L224Q in the PHAGEVO mutational region
BBa_K5061154 Helper phage (HP) M13KO7-ΔgIII-ΔgVI carrying XylS W88C L224Q in the PHAGEVO mutational region
BBa_K5061055 Accessory Plasmid (AP) expressing the M13 gVI and LuxAB from the Pm promoter
BBa_K5061155 Accessory Plasmid (AP) expressing the M13 gIII, M13 gVI and LuxAB from the Pm promoter
BBa_K5061056 Test Plasmid (TP) expressing sfGFP-LVAtag from the Pm promoter
BBa_K5061160 XylS wt expression cassette in EP backbone
BBa_K5061161 XylS K38R L224Q expression cassette in EP backbone
BBa_K5061162 XylS W88C L224Q expression cassette in EP backbone
BBa_K5061163 XylS AI PA expression cassette in EP backbone
BBa_K5061164 XylS AI TPA expression cassette in EP backbone
BBa_K5061165 XylS PHAGEVO-AI v1 PA expression cassette in EP backbone
BBa_K5061166 XylS PHAGEVO-AI v1 TPA expression cassette in EP backbone
BBa_K5061167 XylS PHAGEVO-AI v2 PA expression cassette in EP backbone
BBa_K5061168 XylS PHAGEVO-AI v2 TPA expression cassette in EP backbone
BBa_K5061169 XylS PHAGEVO-AI v3 r3m8 PA expression cassette in EP backbone
BBa_K5061170 XylS PHAGEVO-AI v3 r5m8 PA expression cassette in EP backbone
BBa_K5061171 XylS PHAGEVO-AI v3 r5m10 PA expression cassette in EP backbone
BBa_K5061172 XylS PHAGEVO-AI v3 r5m11 PA expression cassette in EP backbone
BBa_K5061173 XylS PHAGEVO-AI v3 r6m1 TPA expression cassette in EP backbone
BBa_K5061174 XylS PHAGEVO-AI v3 r6m5 TPA expression cassette in EP backbone
BBa_K5061175 XylS PHAGEVO-AI v3 r7m1 PA expression cassette in EP backbone

Basic & Composite Parts for Cas9 & λ Red mediated recombineering

Part Description
BBa_K5061015 ung gRNA
BBa_K5061016 nfi gRNA
BBa_K5061115 ung gRNA transcription unit under the control of the J23119(SpeI) promoter
BBa_K5061116 nfi gRNA transcription unit under the control of the J23119(SpeI) promoter
BBa_K5061215 ung repair DNA
BBa_K5061216 nfi repair DNA

& also

During this project, our team also sequenced the M13KO7 helper phage from New England Biolabs (Cat#N0315S) in order to explain the differences observed upon cloning the HP plasmids listed above. The sequence was deposited in Parts Registry under the Acc number BBa_K5061000.

We have also constructed the E. coli S2060 ∆ungnfi strain and sequenced its genome.

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