User:TChan

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(W 6.14.06)
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==W 6.14.06==
==W 6.14.06==
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1. Miniprep of the 3 BioBricks Plasmids (according to [http://openwetware.org/wiki/Miniprep/Qiagen_kit standard protocol])
 
-
2. Double digest of the 2 R0 (lac promoter) and 2 E0 (gfp) samples
+
<b>Goal:</b> Digest and extract out the E0-gfp and ligate after the R0-promoter, into that pre-cut plasmid.
 +
 
 +
  R0: ---|E--|X---lacprom---|<b>S</b>--|<b>P</b>---
 +
  E0: ---|E--|<b>X</b>---gfp---|S--|<b>P</b>---
 +
  hypothetical ligation: ---|E--
 +
 
 +
1. Miniprepped the 3 BioBricks Plasmids (according to [http://openwetware.org/wiki/Miniprep/Qiagen_kit standard protocol])
 +
2. Double digested the 2 R0 (lac promoter) and 2 E0 (gfp) samples
 +
 
3. Phosphatasing of 2 R0 samples to prevent self-ligation
3. Phosphatasing of 2 R0 samples to prevent self-ligation
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==Tu 6.13.06==
==Tu 6.13.06==
1. Picked Colonies of Existing BioBrick Transformants
1. Picked Colonies of Existing BioBrick Transformants
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*Picked:
+
*Picked and numbered:
   3 colonies of E7,  
   3 colonies of E7,  
   2 colonies of E0,  
   2 colonies of E0,  
-
   2 colonies of R0, and numbered accordingly
+
   2 colonies of R0.
*Grew each colony in 5mL LB + 5uL of 50mg/mL Amp (1:100 dilution).
*Grew each colony in 5mL LB + 5uL of 50mg/mL Amp (1:100 dilution).
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HOMEWORK:
HOMEWORK:
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#Past iGEM Projects Presentation: UCSF
+
1. Past iGEM Projects Presentation: UCSF
-
#Prospective Projects Research:
+
2. Prospective Projects Research:
   Lactobacillus Hijacking
   Lactobacillus Hijacking
   Receptor-based DNA Nanostructure Latch  
   Receptor-based DNA Nanostructure Latch  
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==Contact Info==
==Contact Info==
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/Tiffany Chan
+
  Tiffany Chan
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/Cell: 781 330 1969
+
  Biochemical Sciences
-
/[mailto:chan.tiffany@gmail.com chan.tiffany at gmail.com]
+
  Kirkland '07
 +
  PRISE Summer 2006
 +
  Cell: 781 330 1969
 +
  [mailto:chan.tiffany@gmail.com chan.tiffany at gmail.com]

Revision as of 17:41, 14 June 2006

Contents

W 6.14.06

Goal: Digest and extract out the E0-gfp and ligate after the R0-promoter, into that pre-cut plasmid.

 R0: ---|E--|X---lacprom---|S--|P---
 E0: ---|E--|X---gfp---|S--|P---
 hypothetical ligation: ---|E--

1. Miniprepped the 3 BioBricks Plasmids (according to standard protocol) 2. Double digested the 2 R0 (lac promoter) and 2 E0 (gfp) samples


3. Phosphatasing of 2 R0 samples to prevent self-ligation

4. Ran R0 and E0 samples on gel to separate out the gfp from the E0-plasmid

5. Gel extracted R0 plasmid (~2.1 kb, linearized by digestion) and E0 gfp fragment (~0.9kb)

Tu 6.13.06

1. Picked Colonies of Existing BioBrick Transformants

  • Picked and numbered:
 3 colonies of E7, 
 2 colonies of E0, 
 2 colonies of R0.
  • Grew each colony in 5mL LB + 5uL of 50mg/mL Amp (1:100 dilution).

2. Ran Gel of DNA Nanostructures

  • Ran 2% agarose gel of the nanostructure reaction and the 2 controls for ~40 min at 130V. (NB: Only left four lanes are ours.)
Nanostructure Gel
Nanostructure Gel
 1kb Ladder | Oligo + Scaffold | -Oligo | -Scaffold






M 6.12.06

1. Folded DNA Nanostructures Using Shawn's Pre-designed Oligo Staples and Scaffold ssDNA

oligos + scaffold
-oligos
-scaffold

2. Transformed Bacteria with Existing BioBrick Plasmids

  • Transformed each BioBrick plasmid below into OneShot Top10 competent cells (Invitrogen) according to the following protocol.
 R0010 (lac operon promoter),
 E7104 (T4 promoter + gfp), 
 and E0241 (gfp).

Top10 Transformation

 Thaw cells; aliquot out ~30uL per transformation.
 Add desired amount of plasmid DNA (1 uL in this case?) to cells and leave on ice for 30 min.
 Heat shock at 37&degC for 30 sec in a heating block.  
 Ice for 2 min.
 Shake in 37%deg incubator for 1 hr.
 Pour onto plates with the required antibiotic (carbinomycin in this case); spread with beads or plate spreader.
 Incubate plates in 37%deg incubator overnight.


HOMEWORK: 1. Past iGEM Projects Presentation: UCSF 2. Prospective Projects Research:

 Lactobacillus Hijacking
 Receptor-based DNA Nanostructure Latch 
 Biocryptography w/ DNA Nanostructures
 ?

Contact Info

 Tiffany Chan
 Biochemical Sciences
 Kirkland '07
 PRISE Summer 2006
 Cell: 781 330 1969
 chan.tiffany at gmail.com
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