Adam Crego's Laboratory Notebook: Difference between revisions
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<div style="padding: 10px; width: 750px; border: 5px solid black;"> | |||
{{Template:BrowniGEM2008}} | |||
==02/06/08 '''Transformation of DNA'''== | ==02/06/08 '''Transformation of DNA'''== | ||
Name: control, Part BBa_J45219 | Name: control, Part BBa_J45219 | ||
Line 7: | Line 10: | ||
Parts: (B0032) --> ATF1 (J45014) --> (B0010) --> (B0012) | Parts: (B0032) --> ATF1 (J45014) --> (B0010) --> (B0012) | ||
Library | |||
iGEM 2007 3J iGEM 2007 Parts Kit Plate 3 pSB1AK3 V1009 Available | Library: iGEM 2007, Well: 3J, Plate: iGEM 2007 Parts Kit Plate 3, Plasmid: pSB1AK3, Cell: V1009, Available | ||
====Transformation of Recombinant DNA==== | ====Transformation of Recombinant DNA==== | ||
#) Obtain specific plate from refrigerator. A number and a letter. ex. 3J | |||
#) Set to 8 or 9 microL on pipette | |||
#) Inject distilled water into the well (pipette up and down a few times) and mix. | |||
#) Water should be orange from DNA. | |||
****Only need 4 microL for a transformation (part/date/initials on tube) | ****Only need 4 microL for a transformation (part/date/initials on tube) | ||
Line 21: | Line 25: | ||
====Transformations:==== | ====Transformations:==== | ||
#) If using DNA from Registry, dilute with 9 microL of H20. | |||
#) Label 2 tubes w/ (part/date/name) | |||
#) Add 100 microL competent cells (strain of E. coli : JM109, company Promega, catalog #L2001) in each tube | |||
#) Add 4 microL of Registry DNA into only one tube | |||
#) Place in ice bath for 30 minutes | |||
#) Heat shock at 42 degrees C for 45-60 seconds | |||
#) Place in ice bath for 2 minutes | |||
#) Add 900 microL ice cold antibiotic free LB to cells | |||
#) Incubate at 37 degrees C for 90 minutes in a shaking incubator | |||
#) Dilute as desired, make necessary controls, and plate cells (200 microL per plate) | |||
#) Incubate upside down at 37 degrees C for 18-24 hours, then refrigerate | |||
3 Plates: | ====3 Plates:==== | ||
#) Control: 100 microL LB/cell | |||
#) 2 test plates | |||
1:1 100 micro LB/cells | *1:1 100 micro LB/cells | ||
1:100 1 microL cells to 99 microL H20 | *1:100 1 microL cells to 99 microL H20 |
Latest revision as of 14:51, 6 February 2008
02/06/08 Transformation of DNA
Name: control, Part BBa_J45219
Group 1:
Parts: (B0032) --> ATF1 (J45014) --> (B0010) --> (B0012)
Library: iGEM 2007, Well: 3J, Plate: iGEM 2007 Parts Kit Plate 3, Plasmid: pSB1AK3, Cell: V1009, Available
Transformation of Recombinant DNA
- ) Obtain specific plate from refrigerator. A number and a letter. ex. 3J
- ) Set to 8 or 9 microL on pipette
- ) Inject distilled water into the well (pipette up and down a few times) and mix.
- ) Water should be orange from DNA.
- Only need 4 microL for a transformation (part/date/initials on tube)
• Making lots of DNA - Amplifying using PCR • Save some cells (DNA) in freezer at -80 degrees C
Transformations:
- ) If using DNA from Registry, dilute with 9 microL of H20.
- ) Label 2 tubes w/ (part/date/name)
- ) Add 100 microL competent cells (strain of E. coli : JM109, company Promega, catalog #L2001) in each tube
- ) Add 4 microL of Registry DNA into only one tube
- ) Place in ice bath for 30 minutes
- ) Heat shock at 42 degrees C for 45-60 seconds
- ) Place in ice bath for 2 minutes
- ) Add 900 microL ice cold antibiotic free LB to cells
- ) Incubate at 37 degrees C for 90 minutes in a shaking incubator
- ) Dilute as desired, make necessary controls, and plate cells (200 microL per plate)
- ) Incubate upside down at 37 degrees C for 18-24 hours, then refrigerate
3 Plates:
- ) Control: 100 microL LB/cell
- ) 2 test plates
*1:1 100 micro LB/cells *1:100 1 microL cells to 99 microL H20