Bioware Lab Work

1
Bioware Lab Work Bootcamp!

• Clone two Biobricks from the parts registry and
  combine them in a plasmid.

2
Projects

• Tom and Amanda constitutive pTet promoter and
  ECFP
• Francis and Bob
• Steve and Matt UV inducible promoter and YFP
• Meagan and Erin constitutive pTet promoter and
  GFP (parts E0240 and E0840)

Plasmid Backbone from J04450 CAP resistance and
RFP Biobrick
3
Procedure

• Remove plasmids from registry Transform
• Miniprep, Digest, Ligate, Transform
• Select for non RFP cells that grow on CAP
• 2 overnight cultures to prepare for a
  fluorescence reading
• Confirmation using PCR amplification of Biobrick
  sequequence inside the prefix and suffix.

4
Plates
Meagan and Erin 12M 120 pTet GFP
Transformations 5/26/2010
RFP
Tom and Amanda pTet ECFP Transformations 5/26/
2010
5
Promoter Digest 2 Gel
1 2 3 4 5 6 7 8
9 10
1. 1 kb ladder 2. Constitutive promoter 3. UV
promoter 4. Rx1 Promoter 5. 18C promoter 1 6. 18
C promoter 2 7. RFP 8. Empty 9. Empty 10. Empty
6
Digestions 1 Gel
1. 1 kb ladder 2. ECFP 3. RFP (for backbone) 4.
YFP 5. RFP (for backbone 6. Downstream Part? 7.
RFP (for backbone) 8. 12O GFP 9. 12M GFP 10. RFP
(for backbone)
1 2 3 4 5 6 7 8 9 10
7
Fluorescence Picture
GFP YFP RFP GFP
Exitation 501 512 584 439
Emission 511 529 687 476
8
Possible Directions

• Super metal-resistant cell
• Magnetosomes
• Metal mobilization
• Metal immobilization

9
Metal Resistant Cell
10
Metal Resistant Cell
11
Past Projects

• 2009
• Cornell-Cd sensor-bronze
• Groningen-As accumulation/remediator-gold,
  finalist
• Seoul-Cd As Zn sensor
• Newcastle-Cd sensor/remediator-gold
• Tokyo Tech-Fe oxidation-silver
• Australia-Hg accumulation, ice precipitation-bronz
  e
• Virginia-As accumulation-bronze

• 2008
• UNAM Mexico-Ni Transport-bronze
• METU-Turkey-heavy metal
• sequestration
• Prairieview-Cation transporters-bronze
• 2007
• St. Petersburg-Cu sensor
• Brown-Pb sensor
• MIT-Hg sensor/deactivator
• Prairieview-Fe(II), V(II), Ni(II), S sensor
• Turkey-Metal Transporter
• 2006
• Edinburgh-As sensor
• Latin America-Fe sensor