1010-pBad24-HTNCre
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Cloning of 1010-pBad24-HTNCre
[edit]Purpose: Cloning of 1010-pBad24-HTNCre
Date: 2012-2-28
Notebook:003752
Page:
Clone ID:1010
Experimenters: yingying
Results
[edit]subcloning
[edit]Date:2012-2-28
| band 3 |
band 4 | |
| pTriEx-HTNC (Cre) 298 |
pBad24 545 | |
| digest enzymes
|
Nco1 + Xho1 |
Nco1 + Sal1 |
Colony Screening
[edit]Date: 2012-3-1
Make miniprep and digest with HindIII
Purification
clone 2,3,4,6 may be right
Result intepretation/Summary
[edit]- <br>
Conclusion
[edit]Purification
[edit]11/1/2019
Use Nico21(DE3)pRare2pLysS, 15 ml, induced with 1 mM IPTG, 37 oC 5 hours
cre in Nico21(DE3)pLysN, (from freezer, not freshly transformed) 8L, induce 37 oC 6 hours, 0.15% arabinose., use 50 ml Ni column
| Lane | 1 | MW | Expression | |
| 1 | 1178 | pET28b-scIHF2, soluble |
22.8 | N |
| 2 | 406 | pET28b-Int, soluble |
42.5 | N |
| 3 | 1185 | pDB.His.MBP-Int, soluble |
83.7 | Y |
| 4 | 2640 | pCold-MBP-(Ent-TEV)-scIHF, soluble |
66.9 | N |
| 5 | 2640 | pCold-MBP-(Ent-TEV)-scIHF, inclusion body |
66.9 |
Y |
| 6~12 | 1010 | Cre, |
could be low | |
| 13 | 1178 | pET28b-scIHF2 |
22.8 |
N |
| 14 | 406 | pET28b-Int |
42.5 |
N |
| M | PageRuler Plus |
