M er ed i t h G en t r y

E m m a G o ld s te in

G r a c e H er sh b er g e r

S a m H ig g in s

P ip e r I n g l i s

Ty le r Kn ox

M o l l y Pet ro u s

M a c ken z ie Reet z

M a d iso n S a lo m o n

A m n a S o h a i l

S h a fe r S te l l em a

M r s . Ke l l y B o z o ia n I n s t r u c to r

M r. Pa u l L eg g e

C o n su l t a n t

EARLY MELANOMA

DETECTION

LUMINESCENT

BIOSENSOR

Yeast based malignant

melanoma biosensor

Luminescent cream

Cheap

Efficient, early diagnosis

Readily available

INTENT

Yeast containment

Safe strand of

yeast

Protective gear

Safety precautions

SAFETY

MELANOMA RESEARCH

Figure 2

Figure 2

Figure 3

Figure 2 & 3: Early Stage Melanoma

Source: National Cancer Institute

Figure 1: Melanoma Mortality Rates in U.S. (2000-2004)

Source: National Cancer Institute

MELANOCYTE PATHWAYS RESEARCH

Melanocyte Pathways

Source: National Center for Biotechnology Information

Figure 4

Figure 5

Melanocyte Pathways

Source: Nature Reviews Clinical Oncology

FGFR-1 & FGF2 RESEARCH

Figure 6

Figure 7

Source: Beatson Institute for Cancer Research

Source: Cell Adhesion and Matrix Research

Center, University of Alabama

FGFR-1 & FRS2 RESEARCH

Source: National Center for Biotechnology Information

Source: National Center for Biotechnology

Information

Figure 9

Figure 8

3A General Assembly

Dr. Goebl yeast

Promega

Lab mishaps

PROTOCOL

YEAST BIOSENSOR PHASES & BIOBRICKS

1. Detection of Input (FGF2

Ligand)

2. Activation of Signaling

Pathway

3. Activating the Response

(Output – Luciferase/Renilla

Expression)

FGFR

-1

FRS2

Luciferase & Renilla

YEAST CHASSIS

1

2

3

PHASE ONE: DETECTION OF INPUT

Fibroblast

Growth Factor

Receptor-1

(FGFR-1)

PHASE TWO: ACTIVATION OF SIGNALING PATHWAY

Fibroblast

Growth Factor

Receptor

Substrate 2

(FRS2)

•BBa_K976009: TDH3 Promoter + Yeast Kozak + FGFR-1/FRS2 Complex +TEF1 Terminator

B C D E

A

•FGFR-1 & FRS2 Fusion Protein

Translational Unit BioBrick

DETECTION AND ACTIVATION

PHASE THREE: ACTIVATING RESPONSE

Autonomously

Replicating

Sequence

Binding Factor 1

(ABF1)

PHASE THREE: ACTIVATING RESPONSE

Promoter Control

psbA2 Reference

Dual Luciferase

Assay

A

B

C D

E

F G H

•Linear Sequence of BBa_K654050 – Promoter Control psbA2 Reference Dual

Luciferase Assay

BioBrick Key:

A & E: psbA2 promoter, 5’UTR, and RBS

B: Firefly Luciferase

C &G: T1 terminator from E. coli rrnB

D & H: TE terminator from coliphage T7

F: Renilla luciferase reporter

PHASE THREE: ACTIVATING RESPONSE

C

D

A

B

•Bba_K976008: Yeast ADH1 Promoter + Firefly Luciferase + ADH1 Promoter +

Renilla Luciferase

PHASE THREE: ACTIVATING RESPONSE

Expression Vector Credit: (Kristin T. Chun, Christopher J. Skalnik, and Mark G. Goebl),

Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN

PHASE THREE: ACTIVATING RESPONSE

Yeast ADH1 promoter and pKC3161 Yeast Plasmid Expression Vector

BBa_K976008

failed to

luminesce

Developed new

plasmid and

expression

vector

RESULTS

Dual Luciferase System Growth

Short term:

BBa_K976008

luminesce

Synthesize & transform

FGFR-1 & FRS2 BioBrick

Long term:

Signaling pathway phase

Test FGFR-1 and FGF2

binding

Cream development

NEXT STEPS

FGFR-1 & FGF2 binding

Source: Institut Européen de Chimie et Biologie (IECB)

SPONSORS

Bill Vincent

Dr. Goebl & Dr. Chun

REFERENCES

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