"
Team:University of Lethbridge/Notebook/GeneralLabJune
From 2008.igem.org
Nathan.puhl (Talk | contribs) |
|||
| Line 36: | Line 36: | ||
===June 16 2008=== | ===June 16 2008=== | ||
====Nathan Puhl, Munima, Christa, Sebastian, Roxanne==== | ====Nathan Puhl, Munima, Christa, Sebastian, Roxanne==== | ||
| - | |||
| - | |||
Transformed supercompetent cells with basic biobrick vector pSB1A7 (ampicillin resistance) | Transformed supercompetent cells with basic biobrick vector pSB1A7 (ampicillin resistance) | ||
| - | -50 uL of cells, 0.85 uL 2-mercaptoethanol, 1 uL of plasmid dissolved with 15 uL of ddH2O | + | -50 uL of DH5-alpha cells, 0.85 uL 2-mercaptoethanol, 1 uL of plasmid dissolved with 15 uL of ddH2O |
-30 min on ice | -30 min on ice | ||
-45 s at 42 C | -45 s at 42 C | ||
| Line 49: | Line 47: | ||
====Munima, Christa, Nathan Puhl==== | ====Munima, Christa, Nathan Puhl==== | ||
Checked plates | Checked plates | ||
| - | |||
-Only one colony from transformation, not very good efficiency, don't know why because too many possibilities, most | -Only one colony from transformation, not very good efficiency, don't know why because too many possibilities, most | ||
likely amount of DNA due to inability to quantify plasmid from iGEM plates | likely amount of DNA due to inability to quantify plasmid from iGEM plates | ||
| Line 68: | Line 65: | ||
[[Image:pSB1A7 plasmid.jpg|500 px]] | [[Image:pSB1A7 plasmid.jpg|500 px]] | ||
| + | |||
| + | plasmid is ~15 ng/uL | ||
| Line 77: | Line 76: | ||
===June 25, 2008=== | ===June 25, 2008=== | ||
====Nathan Puhl, Sebastian, Alix==== | ====Nathan Puhl, Sebastian, Alix==== | ||
| - | Subcultured | + | Subcultured BBa_I13522 into liquid LB + amp for plasmid mini prep |
| Line 85: | Line 84: | ||
Plasmid mini-prepped GFP complete (BBa_I13522). | Plasmid mini-prepped GFP complete (BBa_I13522). | ||
| + | |||
| + | ===June 27, 2008=== | ||
| + | ====Nathan Puhl, Alix, Munima==== | ||
| + | No colonies on any plates. Will try again next week. | ||
| + | |||
| + | ===July 1, 2008=== | ||
| + | ====Nathan Puhl, Andrew==== | ||
| + | Made 500 mL of LB agar + amp and 500 mL of Liquid LB | ||
| + | |||
| + | ===July 2, 2008=== | ||
| + | ====Nathan, ALix, Sebastian, Munima, Roxanne, Crista==== | ||
| + | |||
| + | Trasformed BBa_J24679( RBS + LacI), BBa_P0440 (RBS+TetR+T10+T12), leftover DNA from Double T (June 26, 2008), and 1 uL of pSB1A7 plasmid (June 18). | ||
| + | |||
| + | Protocol changes | ||
| + | -3 uL of DNA | ||
| + | -spin down 200 uL of cells and resuspend in 100 uL of LB; plate on LB + amp | ||
Revision as of 00:52, 3 July 2008
Contents |
June 6 2008
Sebastian, John and Roxanne
Prepared 1L of semi-solid media following the procedure found on OpenWetWare.
-10g peptone (substituted for tryptone) -10g Agar -10g NaCl -5g Yeast Extract
Stored media in fridge.
June 10 2008
Christa, Munima, Roxanne, and Sebastian
Prepared 1L of liquid media following the procedure found on OpenWetWare.
-10g peptone (substituted for tryptone) -10g NaCl -5g Yeast Extract
Poured 36 culture test tubes, and the remainder was left in a 1L Erlenmeyer flask. Stored both in fridge.
Christa
Made an inventory of iGEM 2007 parts in Wieden -80 freezer inventory.xls
Roxanne
Defrosted the -20 freezer in the teaching lab for iGEM use (with a little help from my friends!)
June 11 2008
Sebastian, Munima, Roxanne, Christa
Poured 8 minimal media (labeled control - with blue sharpie) plates and 14 amp plates (labeled amp - with red sharpie) stored in the 4 C fridge. Amp concentration is always 50ug/mL.
June 16 2008
Nathan Puhl, Munima, Christa, Sebastian, Roxanne
Transformed supercompetent cells with basic biobrick vector pSB1A7 (ampicillin resistance)
-50 uL of DH5-alpha cells, 0.85 uL 2-mercaptoethanol, 1 uL of plasmid dissolved with 15 uL of ddH2O -30 min on ice -45 s at 42 C -2 min on ice -Add 0.9 mL of LB shaker incubate at 225 RPM and 37 C
June 17 2008
Munima, Christa, Nathan Puhl
Checked plates
-Only one colony from transformation, not very good efficiency, don't know why because too many possibilities, most likely amount of DNA due to inability to quantify plasmid from iGEM plates -Subcultured colony in liquid LB + amp -Plate 200 uL on LB + amp at 37 C overnight
June 18 2008
Munima, Christa, Alix, Nathan Puhl
Made glycerol stock of pSB1A7 transformed E. coli
Extracted plasmid from transformed E. coli using the Eppendorf FastPlasmid minikit and stored 4 aliquots of 25 uL in the -20 C freezer.
June 19 2008
Nathan Puhl, Alix, Munima, Christa, Roxanne
Ran plasmids on 1% agarose gel with High range ladder
plasmid is ~15 ng/uL
June 24 2008
Nathan Puhl, Alix
Streaked BBa_I13522 (TetR repressed GFP) onto LB + amp from last year's glycerol stock.
June 25, 2008
Nathan Puhl, Sebastian, Alix
Subcultured BBa_I13522 into liquid LB + amp for plasmid mini prep
June 26, 2008
Nathan Puhl, Sebastian, Alix
Transformed RFP complete (BBa_J5526), pLACI-|TetR (BBa_I730002), Double T (BBa_B0015)
Plasmid mini-prepped GFP complete (BBa_I13522).
June 27, 2008
Nathan Puhl, Alix, Munima
No colonies on any plates. Will try again next week.
July 1, 2008
Nathan Puhl, Andrew
Made 500 mL of LB agar + amp and 500 mL of Liquid LB
July 2, 2008
Nathan, ALix, Sebastian, Munima, Roxanne, Crista
Trasformed BBa_J24679( RBS + LacI), BBa_P0440 (RBS+TetR+T10+T12), leftover DNA from Double T (June 26, 2008), and 1 uL of pSB1A7 plasmid (June 18).
Protocol changes
-3 uL of DNA -spin down 200 uL of cells and resuspend in 100 uL of LB; plate on LB + amp
