User:TChan: Difference between revisions
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==W 6.14.06== | ==W 6.14.06== | ||
2. Double | <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 | ||
*Picked: | *Picked and numbered: | ||
3 colonies of E7, | 3 colonies of E7, | ||
2 colonies of E0, | 2 colonies of E0, | ||
2 colonies of R0 | 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: | ||
1. Past iGEM Projects Presentation: UCSF | |||
2. Prospective Projects Research: | |||
Lactobacillus Hijacking | Lactobacillus Hijacking | ||
Receptor-based DNA Nanostructure Latch | Receptor-based DNA Nanostructure Latch | ||
Line 66: | Line 73: | ||
==Contact Info== | ==Contact Info== | ||
Tiffany Chan | |||
Biochemical Sciences | |||
Kirkland '07 | |||
PRISE Summer 2006 | |||
Cell: 781 330 1969 | |||
[mailto:chan.tiffany@gmail.com chan.tiffany at gmail.com] |
Revision as of 14:41, 14 June 2006
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.)
1kb Ladder | Oligo + Scaffold | -Oligo | -Scaffold
M 6.12.06
1. Folded DNA Nanostructures Using Shawn's Pre-designed Oligo Staples and Scaffold ssDNA
- Three reactions below were mixed according to given protocol.
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°C 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