Imperial/Wet Lab/Protocols/CE1.3
From 2007.igem.org
m |
m |
||
| (2 intermediate revisions not shown) | |||
| Line 1: | Line 1: | ||
{{Template:IC07navmenu}} | {{Template:IC07navmenu}} | ||
| + | <br clear="all"> | ||
__NOTOC__ | __NOTOC__ | ||
| Line 5: | Line 6: | ||
==Aims== | ==Aims== | ||
| - | + | Proper experimental amounts for reaction mixtures of S30 E.coli cell extract from Promega. | |
==Equipment== | ==Equipment== | ||
| Line 27: | Line 28: | ||
#*20ul of S30 Premix without amino acids | #*20ul of S30 Premix without amino acids | ||
This makes up a total of 40ul. | This makes up a total of 40ul. | ||
| - | #Add the appropriate amount of DNA to the reaction mixture. (For our experiments we use 2ug and 4ug worth of DNA) | + | #Add the appropriate amount of DNA to the reaction mixture. (For our experiments we use 2ug and 4ug worth of DNA.) |
#Top up the mixture to 60ul with nuclease free water. | #Top up the mixture to 60ul with nuclease free water. | ||
Latest revision as of 02:13, 27 October 2007
Using the commercial E. Coli cell extract
Aims
Proper experimental amounts for reaction mixtures of S30 E.coli cell extract from Promega.
Equipment
- Eppendorf Tubes
- Gilson p20,p200,p1000
Reagents
- Commercial S30 E.coli extract. Including:
- 175µl Amino Acid Mixture Minus Cysteine, 1mM
- 175µl Amino Acid Mixture Minus Methionine, 1mM
- 175µl Amino Acid Mixture Minus Leucine, 1mM
- 450µl S30 Extract, Circular (3 × 150µl)
- 750µl S30 Premix Without Amino Acids
- Nuclease Free water x1ml
- DNA from midiprep/maxiprep
Protocols
- From the commercially available kit, one reaction mixture comprises of:
- 2.5ul of two different types of amino acid mixtures (total volume of 5ul)
- 15ul of S30 Extract, Circular
- 20ul of S30 Premix without amino acids
This makes up a total of 40ul.
- Add the appropriate amount of DNA to the reaction mixture. (For our experiments we use 2ug and 4ug worth of DNA.)
- Top up the mixture to 60ul with nuclease free water.
