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| - | == 6-15-2009 ==
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| - | Lab: LV, SH, CZ
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| - | '''SH''':
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| - | * [[Team:Heidelberg/n4tr1um#Isolation of plasmid DNA by alkaline lysis (mini and maxiprep)|Miniprep]] of GFP template plasmid, pcDNA5/FRT
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| - | {| class="wikitable" border="1"
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| - | |-
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| - | ! Nr
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| - | ! pcDNA5/FRT
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| - | ! GFP
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| - | |-
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| - | | 1
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| - | | 4,7
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| - | | 39,7
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| - | |-
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| - | | 2
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| - | | 5,9
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| - | | 14,9
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| - | |-
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| - | | 3
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| - | | 3,7
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| - | | 3,8
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| - | |-
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| - | | 4
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| - | | 5,0
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| - | | 7,8
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| - | |-
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| - | |}
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| - | * [[Team:Heidelberg/n4tr1um#Isolation of plasmid DNA by alkaline lysis (mini and maxiprep)|Maxiprep]] pcDNA/FRT 188.7ng/µL
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| - |
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| - | '''LV''':
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| - | * Extraction of CMV promoter from 2008 distribution
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| - | * [[Team:Heidelberg/n4tr1um# Transformation of Bacteria|Transformation]] of DH5a cell with CMV promoter
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| - | * portzughtr
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| - |
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| - | == 6-16-2009 ==
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| - |
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| - | Lab: LV, SH, CZ
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| - |
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| - | * DNA synthesis (JeT, cFos, Min)
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| - | * Purification of cDNA via 2% / 3% agarosegel
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| - | => Only bands at ca. 50 Bp => no successful amplification
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| - |
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| - | * [[Team:Heidelberg/n4tr1um#Site-directed mutagenesis|Site directed mutagenesis]] of pcDNA5/FRT and mcherry
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| - | * Dpn1 digestion
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| - | * Transformation of DH5a
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| - | == 6-17-2009 ==
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| - | Lab: LV, SH
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| - | * No transformations could be observed
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| - | * Replace Phsuion stocks to Phusion Master Mix from Nathan
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| - | * Repeat DNA synthesis with a different protocol: 1:10 - 1:1000 dilution of Oligos, 95° 5', 7* [95° 45'' 58° 45'' 72° 1'], add 1:10 diluted primers, 95° 5' 25* same temperatures
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| - | * Annealing successful for JeT and Min
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| - | Annealing might not have been successful with Fos beacuse it contains a very repetetive proximal promoter.
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| - | * gel purfifcation of JeT and Min
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| - | * Repeat PCR of pcDNA5/FRT and mcherry with lower annealing temperature (58°C) and fresh Polymerase: Follow PCR protcol from Strategene site directed mutagenesis kit.
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| - | mcherry PCR might not have been successful because we used a template we didn't make ourselves - does it contain TE?
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| - | * PCR worked for pcDNA5FRT but not for mcherry
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| - | * DpnI digest of pcDNA5FRT 1,3,4,6
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| - |
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| - | == 6-18-2009 ==
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| - |
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| - | Lab: LV, SH
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| - | * DNA synthesis for Fos (proximal) and Fos (core)
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| - | * Transformation of DH5a with pcDNA/FRT ΔPstI
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