Team:Groningen/Notebook/27 July 2009
From 2009.igem.org
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PCR 2 was add on gel again as noted below at accumulation. | PCR 2 was add on gel again as noted below at accumulation. | ||
- | + | PCR 1 was | |
'''GlpF PCR1''' | '''GlpF PCR1''' | ||
{| | {| | ||
Line 76: | Line 76: | ||
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+ | A 2% agarose gel was used, a band of 105 bp was expected. | ||
[[Image:F102471_2009-07-27_16hr_21min_GlpFPCR1_noted.JPG]] | [[Image:F102471_2009-07-27_16hr_21min_GlpFPCR1_noted.JPG]] | ||
+ | |||
+ | The band in row 2 was cut and purified using the ... kit. | ||
+ | |||
+ | The purified PCR1 and PCR2 were used to run PCR3 | ||
+ | |||
+ | |||
+ | '''GlpF PCR3''' | ||
+ | {| | ||
+ | | | ||
+ | <!--Tabel 1 hier--> | ||
+ | |||
+ | {| border="1" | ||
+ | |+ '''PCR1.1''' | ||
+ | ! Component !! amount | ||
+ | |- | ||
+ | ! 2x Phusion MM | ||
+ | | 12.5 uL | ||
+ | |- | ||
+ | ! GlpF PCR1 | ||
+ | | 6 uL | ||
+ | |- | ||
+ | ! GlpF PCR2 | ||
+ | | 6 uL | ||
+ | |} | ||
+ | |||
+ | |width="10"| | ||
+ | | | ||
+ | <!--Tabel 2 hier--> | ||
+ | {| | ||
+ | ! GlpF PCR1 program | ||
+ | ! Temperature | ||
+ | ! Time | ||
+ | |- | ||
+ | |Denaturing | ||
+ | |98° | ||
+ | |2.00 min | ||
+ | |- | ||
+ | |Start Cycles 30X | ||
+ | |- | ||
+ | |Denaturing | ||
+ | |98° | ||
+ | |10 sec | ||
+ | |- | ||
+ | |Annealing | ||
+ | |60° | ||
+ | |10 sec | ||
+ | |- | ||
+ | |Elongation | ||
+ | |72° | ||
+ | |30 sec | ||
+ | |- | ||
+ | | | ||
+ | |End cycles | ||
+ | |- | ||
+ | |Final elongation | ||
+ | |72° | ||
+ | |5 min | ||
+ | |- | ||
+ | |Hold | ||
+ | |4° | ||
+ | |Forever | ||
+ | |} | ||
+ | |width="10"| | ||
+ | | | ||
+ | |} | ||
===Metal Accumulation=== | ===Metal Accumulation=== |
Revision as of 16:04, 27 July 2009
[http://2009.igem.org/Team:Groningen http://2009.igem.org/wiki/images/f/f1/Igemhomelogo.png]
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Wet
GVP Cluster
Transporters
GlpF PCR 2 was add on gel again as noted below at accumulation.
PCR 1 was GlpF PCR1
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A 2% agarose gel was used, a band of 105 bp was expected.
The band in row 2 was cut and purified using the ... kit.
The purified PCR1 and PCR2 were used to run PCR3
GlpF PCR3
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Metal Accumulation
Gel with ArsR PCR((24/07/09)
ArsR/GlpF gel extraction
- - Used zymo research Zymoclean TM Gel DNA Recovery Kit to extract ArsR, GlpF PCR2 and GlpF FullLenght from gel (cut from gel as shown in figure above).
GlpF PCR1
12,5 uL | Phusion |
1 uL | GlpF FW primer |
1 uL | GlpF RevMut primer |
0,25 uL | GlpF Full Lenght DNA (extracted as noted above)(not in tube2) |
10,25 uL | MQ |
Vectors
Dry
Finished RPU computations!
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