User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2013/04/22

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(Autocreate 2013/04/22 Entry for User:Behzad_Damadzadeh/Notebook/PcTF_Subcloning_in_E-coli)
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'''List title'''
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'''Building a Reporter Gene to Determine Chromatin Protein Function (Ligation into the mammalian vector)'''
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# List items
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*Ligation of construct KAH182-KAH201 and mammalian vector V0200 and cloning in DH5α-T..
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KAH182 = BBa_S04739 and KAH201 = BBa_S04745 and V0200 = BBa_J176121. 
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*Assembly strategy: The backbone KAH201(S/P)/3403 , the insert is KAH182(X/P)/1675.
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'''Digests''' <br>
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[[Image:KAH201-KAH182 Ligation.jpg|thumb|400px| (1,2) KAH182, 1675 bp, Cut with Xba1 and Pst1, (3,4) KAH201, 3403 bp, Cut with Spe1 and Pst1]]
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{| {{table}} cellspacing="3"  width=350px
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| Plasmid DNA || 25.00 μl
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|-
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| NotI  || 1.0 μl
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|-
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| SpeI || 1.0 μl
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|-
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| 10x FastDigest buffer + green loading dye || 3.00 μl
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|-
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| dH<sub>2</sub>O || 0.00 μl
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|-
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| &nbsp; || 30.0 μl total
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|-
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| Incubate at 37°C for 10 minutes.
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|}

Revision as of 18:38, 22 April 2013

PcTF Subcloning in E. coli Main project page
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Date

Building a Reporter Gene to Determine Chromatin Protein Function (Ligation into the mammalian vector)


  • Ligation of construct KAH182-KAH201 and mammalian vector V0200 and cloning in DH5α-T..

KAH182 = BBa_S04739 and KAH201 = BBa_S04745 and V0200 = BBa_J176121.

  • Assembly strategy: The backbone KAH201(S/P)/3403 , the insert is KAH182(X/P)/1675.


Digests


(1,2) KAH182, 1675 bp, Cut with Xba1 and Pst1, (3,4) KAH201, 3403 bp, Cut with Spe1 and Pst1
(1,2) KAH182, 1675 bp, Cut with Xba1 and Pst1, (3,4) KAH201, 3403 bp, Cut with Spe1 and Pst1
Plasmid DNA 25.00 μl
NotI 1.0 μl
SpeI 1.0 μl
10x FastDigest buffer + green loading dye 3.00 μl
dH2O 0.00 μl
  30.0 μl total
Incubate at 37°C for 10 minutes.
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