1656-pCold-GST-(3c)-KEAP1
Cloning of 1656-pCold-GST-(3c)-KEAP1
[edit]Purpose: Cloning of 1656-pCold-GST-(3c)-KEAP1
Date: 2013-2-5---2-13-2-7
Notebook:0
Page:
Clone ID:1656
Experimenters: yingying
Results
[edit]cloning
[edit]
subclone from 1625 pT7A3-MBP-(3c)-KEAP1 using Nde1 + Xho1 to 1629 pCold-GST-(3C)
Colony Screening
[edit]Date: 2013-2-7
Make miniprep and digest with Nde1 + Xho1
Result intepretation/Summary
[edit]- <br>
Conclusion
[edit]Protein Expression
[edit]Protein Expression
[edit]2013/2/11
| Lane |
Name |
Clone ID |
Host |
Induction |
Purification |
| M |
Invitrogen SeeBlue plus2 |
||||
| 1 |
pT7A3-MBP-(3c)-KEAP1 |
1625 |
4 ml, BL21(DE3)Star, pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads |
| 2 |
pT7A3-MBP-(3c)-KEAP1 |
1625 |
4 ml, BL21(DE3)Star, pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads |
| 3 |
pCold-GST-(3c)-KEAP1 |
1656 |
4 ml, BL21(DE3)Star, pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads (wrong, should use Glutathione agarose beads) |
| 4 |
pCold-GST-(3c)-KEAP1 |
1656 |
4 ml, BL21(DE3)Star, pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads (wrong, should use Glutathione agarose beads) |
| 5 |
pET28-cold-NRF2 |
1626 |
4 ml, BL21(DE3)Star, pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads |
| 6 |
pT7A3-MBP-i3C |
1554 |
4 ml,NiCO21(DE3), pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads |
| 7 |
pT7A3-6His-GST-i3C |
1601 |
4 ml,NiCO21(DE3), pRare2-pLysS |
0.2 mM IPTG 25 oC o/n |
Ni beads |
| 8 |
Protein expression of KEAP1 clones
[edit]| clone ID |
expression |
ge image |
|
| 1625,pT7A3-MBP-(3c)-KEAP1 |
yes |
1625 1626 1656 |
|
| 1656,pCold-GST-(3c)-KEAP1 |
need to use GST beads |
1625 1626 1656 |
|
| 1676,pCold-MBP-(3C)-KEAP1 |
yes |
1661 1674 1683 |
|
| 1701,pMAL-c5E-KEAP1 |
yes |
||
| 1720,pT7A3-MBP-(E3C)-Keap1 |
no |
626 1717 1719 1720 |
|