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| | * Amanda Ispas: Buffers, media (and quality control), preparation of BioBrick registry submissions | | * Amanda Ispas: Buffers, media (and quality control), preparation of BioBrick registry submissions |
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| - | ==Daily Lab Notebook==
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| - | {|{{table}} width="800"
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| - | |style="background-color: #cdde95;" align="center"|
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| - | <!-- ## START calendar column -->
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| - | <LNCalendar></LNCalendar>
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| - | <!-- ## END calendar column -->
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| - | |align="center" style="background-color: #e5edc8;" |
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| - | <!-- ## START search column: Place your logo here. Try keep it below 200px in width and 150px in height. ## -->
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| - | [[Image:owwnotebook_icon.png|128px]] <sitesearch>title=Search this Project</sitesearch>
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| - | <!-- ## END search column ## -->
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| - | |-
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| - | |colspan="2" style="background-color: #F2F2F2;" align="right"|[[{{FULLPAGENAME}}/Entry_Base|Customize your entry pages]] [[Help:Notebook/Project_Base/Customize_entry_page|<html><img src="/images/a/aa/Help.png" border="0" /></html>]]
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| - | |colspan="2"|
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| - | ==Project Description/Abstract==
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| - |
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| - | [[#THAT| '''Team Chimeric Reporter:''']]<br>
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| - | * Hyder Hussain
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| - | * Abhinav Markus
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| - | * Ryan Muller
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| - | * Ethan Ward
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| - | [[#THIS| '''Team Cell Surface Reporter:''']]<br>
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| - | * Nisarg Patel
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| - | * Ryan Muller
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| - | * Rohit Rajan
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| - | * Ellen Qin
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| - | * Melinda Jenner
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| - | * Joe Barth
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| - |
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| - | ==BioBricks==
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| - |
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| - | '''Magainin Reporter'''
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| - | *Magainin
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| - | *Magainin + 2xGS Linker + His-Tag
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| - | *Bgal fragment (a-1)
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| - | *Bgal fragment (a-4)
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| - | *Bgal fragment (w)
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| - | *Bgal fragment (1-w)
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| - | *Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (a-1)
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| - | *Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (a-4)
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| - | *Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (w)
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| - | *Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (1-w)
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| - | *His-Tag + 10aa GS Linker + Bgal fragment (a-1)
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| - | *His-Tag + 10aa GS Linker + Bgal fragment (a-4)
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| - | *13. His-Tag + 10aa GS Linker + Bgal fragment (w)
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| - | *14. His-Tag + 10aa GS Linker + Bgal fragment (1-w)
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| - |
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| - | ==Notes==
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| - |
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| - | ===6-7-12===
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| - | * Transformation (LSE)
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| - | :* Transformed DNA:
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| - | ::* lacZ (well 4:12G, I732019)
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| - | ::* p + lacO (well 1:6G, R0011)
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| - | :* Cells: neb10beta (donated)
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| - | :* Protocol from: http://www.neb.com/nebecomm/products/productc3019.asp
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| - | :* Controls: puc19, no DNA (8 plates)
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| - |
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| - | ===6-8-12===
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| - | * Transformation results:
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| - | :* puc19: growth
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| - | :* negative control: no growth
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| - | :* lacZ, lacO: possible small colonies
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| - | * liquid culture in amp media (100 ug / ml):
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| - | :* no growth of lacZ, lacO
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| - | :* growth of puc19
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| - |
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| - | ===6-12-12===
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| - | *DH5a Competent Cell Prep
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| - | :*Streak plated cells on LB no amp plate, let grow overnight
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| - |
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| - | ===6-13-12===
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| - | * Transformation (LSE)
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| - | :* Transformed DNA:
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| - | ::* lacZ (well 4:12G, I732019)
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| - | ::* p + lacO (well 1:6G, R0011)
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| - | :* Cells: DH5 alpha (donated)
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| - | :* Protocol from: http://openwetware.org/wiki/Haynes:Assembly101 (30 minute transformation)
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| - | :* Controls: puc19, no DNA (8 plates)
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| - | * Transformation (istb4, Abhi)
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| - | :* Transformed DNA:
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| - | :* Cells:
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| - | :* Protocol from:
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| - | :* Controls:
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| - | *DH5a Chemically Competent cell prep
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| - | :*Grew 2 seed colonies from streak plate in LB no amp
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| - | :*Grew controls to test for contamination
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| - | ::*Both Seed colonies grew, no contamination present
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| - |
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| - | ===6-14-12===
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| - | *Competent cell prep
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| - | :*Prepared CaCl<sub>2</sub> buffer solution and CaCl<sub>2</sub> glycerol buffer solution
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| - | :*Grew seed colony in 400mL LB no amp
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| - |
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| - | ===6-15-12===
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| - | *Competent cell prep
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| - | :*Centrifuged falcon test tubes containing liquid colonies
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| - | :*Resuspended in CaCl<sub>2</sub> buffer solution and incubated for 15 mins
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| - | :*Centrifuged and resuspended in CaCl<sub>2</sub> glycerol buffer solution
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| - | :*Chilled overnight
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| - |
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| - | ===6-16-12===
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| - | *Competent cell prep
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| - | :*Aliquotted 200uL into test tubes
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| - | :*Stored in -80C
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| - |
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| - | ===6-17-12===
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| - | :*Streak plated prepared competent cells on LB no amp plate
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| - | ::*Colonies observed
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| - |
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| - | ===6-19-12===
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| - | *Transformation (LSE)
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| - | :*Transformed DNA:
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| - | ::*T7 promoter BBa_I712074
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| - | ::*Constitutive promoter BBa_J23102
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| - | :* Cells: DH5 alpha (donated)
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| - | :* Protocol from: http://tools.invitrogen.com/content/sfs/manuals/subcloningefficiencydh5alpha_man.pdf (30 minute transformation)
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| - | :* Controls: puc19, no DNA
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| - | :* Plated 2 copies of each (100 ul, 250 ul) on LB amp plates.
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| - | * Made 50 LB Amp plates.
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| - |
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| - | ===6-20-12===
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| - | * Plated negative control on LB Amp plate
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| - | * Liquid cultures of T7 promoter and constitutive promoter
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| - | *Transformation (LSE)
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| - | :*Transformed DNA:
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| - | ::*RBS (well 1:1H BBa_B0030)
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| - | ::*TetR GFP (well 2:8A Part:BBa_I13522)
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| - | :* Cells: DH5 alpha (donated)
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| - | :* Protocol from: http://tools.invitrogen.com/content/sfs/manuals/subcloningefficiencydh5alpha_man.pdf (30 minute transformation)
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| - | :* Controls: puc19, no DNA
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| - |
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| - | ===6-21-12===
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| - | *Made Liquid Cultures of E.coli transformed with RBS B0030
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| - | *Made Liquid Cultures of E.coli transformed with TetR GFP
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| - | *miniprepped and nanodropped T7 promoter BBa_I712074 and Constitutive promoter BBa_J23102 liquid cultures
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| - | * liquid cultures:
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| - | :* RBS1
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| - | :* RBS2 (duplicate_
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| - | :* GFP1
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| - | :* puc19
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| - | :* negative controls
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| - | :* 5 ml LB amp
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| - | :* overnight cultures
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| - | * replated GFP1 & 2 (duplicates)
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| - | *Nanodropped plasmid DNA samples
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| - | :*Constitutive promoter 1: __ng/uL
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| - | :*Constitutive promoter 2: __ng/uL
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| - | :*T7 promoter 1: __ng/uL
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| - | :*T7 promoter 2: __ng/uL
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| - |
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| - | ===6-22-12===
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| - | :*Miniprepped liquid cultures: RBS (well 1:1H BBa_B0030) and TetR GFP (well 2:8A Part:BBa_I13522)
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| - |
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| - | ===6-26-12===
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| - | * Transformation:
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| - | :* Transformed DNA:
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| - | ::* double terminator (B0017, 2:6K)
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| - | ::* T7 RNA polymerase (I715038, 2:15C)
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| - | ::* puc19, negative control
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| - | :* Protocol from: http://tools.invitrogen.com/content/sfs/manuals/subcloningefficiencydh5alpha_man.pdf (30 minute transformation)
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| - | :* Cells: dh5a
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| - |
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| - | ===6-27-12===
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| - | * 6-26 transformation results:
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| - | :* Controls correct
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| - | :* 2x terminator: ~19 colonies
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| - | :* RNA pol: 1 colony
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| - | * Liquid cultures including controls
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| - |
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| - | ===6-28-12===
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| - | *Miniprepped double terminator (B0017, 2:6K) and T7 RNA polymerase (I715038, 2:15C) liquid cultures
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| - |
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| - | ===7-2-12===
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| - | *Cleaned up liquid waste
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| - | *Made SOB media
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| - | *Finalized oligos for magainin construct
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| - |
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| - | ===7-3-12===
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| - | *Autoclaved SOB media
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| - | *Added glucose to make SOC media
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| - | *Nanodropped double terminator (B0017, 2:6K) [DT1: 24.5, DT2: 29.6] and T7 RNA polymerase (I715038, 2:15C) [P1: 64.6, P2: 55.3] liquid cultures
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| - | |-
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| - | |colspan="2" style="background-color: #F2F2F2;"|
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| - | |}
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| - |
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| - | __NOTOC__
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| | ==BioBricks== | | ==BioBricks== |
