Difference between revisions of "Team:Sydney Australia/Parts"

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin

E. Coli (Ecotin Tag)

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We submitted this DNA part to the registry, for other teams to use in the future

N/A – Winsulin is self designed from literature and mammalian insulins

GOLD

PART NAME / NO.

SOURCE

KEY FEATURES

WHAT IS IT GOLD?

YncM-Winsulin

Bacillus subtilis (YncM secretion tag)

Novel short chain insulin analogue (Winsulin) with a 12AA linker
(-QRGGGSGGGQRR-)

YncM secretion tag to translocate protein into media

N-terminal 6x His Tag purification

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

We demonstrated that it works:

· We tested it in insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, proving its bioactivity

· We showed via an ELISA assay that whilst the cell lysate did not contain Winsulin, the media in which the cells were growing did. Therefore the YncM secretion tag worked as expected

The results for these assays can be found here

Ecotin-Proinsulin

E. Coli (Ecotin tag)

Human proinsulin

N-terminal Ecotin tag for targeting proteins to the periplasm

N-terminal 6x His tag

4x GGS flexible linker

trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

completing their sequence

Adding a periplasmic transporter tag (for more efficient folding in an oxidative environment)

Adding an efficient ribosomal binding site

Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

Human (Human proinsulin)

Cytoplasmic-Proinsulin BBa_K2417001

Human (Human proinsulin)

Human proinsulin

N-terminal 6x His tag

4x GGS flexible linker

Trypsin protease cleavage site for His-tag and C-peptide cleavage

Extended ribosomal binding site

We improved previous parts:

BBa_M39904 and BBa_M1877 by:

· completing their sequence

· Adding an efficient ribosomal binding site

· Adding an N-terminal His tag for purification

We demonstrated that it works:

· By testing it insulin-sensitive cell lines and saw an increase in glycogen synthesis and glucose oxidation above basal levels, therefore proving its bioactivity

The results for these assays can be found here

SILVER

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT SILVER?

Cytoplasmic-Winsulin

N/A – Winsulin is self designed from literature and mammalian insulins

Novel single chain insulin analogue (Winsulin) with a 12AA linker (QRGGGSGGGQRR)

N-terminal 6x His Tag

4x GSS flexible linker

TEV protease cleavage site

Extended ribosome binding site

· We validated our parts by showing that they work in producing insulin. We performed an ELISA assay testing for properly folded insulin, and detected the presence of Cytoplasmic Winsulin.

The results for the assay can be found here

BRONZE

PART NAME / NO.

SOURCE

KEY FEATURES

WHY IS IT BRONZE?

Ecotin-Winsulin