931 pET28b-PAK16D ligase: Difference between revisions
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Latest revision as of 12:46, 24 September 2018
--Paul.xu (talk) 15:29, 21 December 2017 (EST)
12/27/2017
938 digestion with Nde1-Xho1, Fastdigest Buffer
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8,9 | 10,11 | 13 |
| 938 PNGase_F | 939 Tte-ExoIII |
942 VVN |
3122 CRM197 |
931, 1, PAK16D ligase |
931, 2, PAK16D ligase |
1855 yPolyA-Pol |
1540 | 2046 | 2046 |
| Nde1-Xho1 | Nde1-Xho1 |
Nco1-Xho1 |
Nde1-Xho1 |
Nde1-HindIII |
HindIII-Xho1 | Nde1-Xho1 | Nde1-Xho1 | Nde1-Xho1 | Nco1-Xho1 |
| pJet1.2 | pJet1.2 |
pJet1.2 |
pJet1.2 |
pJet1.2 |
pJet1.2 |
pEC2- |
pET30b | pET28b | pET28b |
| 1145 |
780 |
716 |
1798 |
1174 |
986 |
1713 | there is no Xho1 site | 1377 | ~1377 |
Date:4/5/2018
[edit]Subclone of 931
Vector:1987pT7A3
Insert:
F1:1174 bp
F2:986 bp
F1:Nde1+HindIII
F2:HindIII+Xho1
Date:5/29/2018
[edit]Subclone of 931
Vector is 1987pT7A3
Miniprep digestion of 984_931
| # | Lane | Enzymes | Size | Note |
| 984 | 2-5 | Xba1+Nhe1 | 957bp | #6,7,,8 seems good |
| 931 | 1,6-8 | HindIII+Xho1 | 986bp | #5 seems good |
Digest with HindIII +Xho1 and Nde1+ HindIII, Nde1 + Xho1
using parental vector as control. Need to have marker
not good
Date:7/10/2018
[edit]931-1 subclone
931 subclone
Miniprep digestion of 931-1
931 miniprep digestion 1
931-1 miniprep digestion
Nde1+HindIII
931subclone
TE as EB buffer
multiple bands in gel, not good, can not cut out the vector.
Date:8/17/2018
[edit]931-1 miniprep
vector:pET28b
insert:931-1 in 1727 vector
931 Subclone
vector is 931-1 in pET28b
HindIII+Xho1
Miniprep digestion of 931
931,#4 digestion
| Lane | enzyme | size |
| 1 | Nde1+HindIII | 1174 |
| 2 | HindIII+Xho1 | 986 |
| 3 | HindIII+Xho1 | 986 |
| 4 | Nde1+Xho1 | 2267 |
Junk fragment , from hindIII+Xho1
Date:9/20/2018
[edit]931 PCR
931 vector digestion
931 PCR Screening
