MS 5 26 2017
Prepare LB media (20L).
Spin down 1708 HTNCre, wash with buffer A, break down through homogenizer, collect supernatant, load onto Ni column.
Troubleshoot with printer, load paper.
Deal with order 4868.
Small scale expression screening of 483 Tth Polymerase /1464 T7 DNA Ligase / 1946 TdT bovine / 1497 ToughTaq. Run SDS-PAGE gel.
| number |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
11 |
12 |
| sample |
483 P |
1464 P |
1946 P |
1497 P |
MW |
483 S |
1464 S |
1946 S |
1497 S |
483 U |
1464 U |
1946 U |
| uL |
15 |
15 |
15 |
15 |
10 |
15 |
15 |
15 |
15 |
15 |
15 |
15 |
S: use lysis buffer S to break down cell, spin down and collect supernatant.
P: use lysis buffer P to wash sediment, sse blunted needle to break pellet, spin down and collect supernatant.
U: Uninduced sample. S and P induced with IPTG.
Lane 6~9 have bands. It means there is protein expression occur in both induced sample.
Lane 2 is brighter than lane 7, both of the two lanes load 1464 T7 DNA Ligase. It means there is more protein in the sediment than the supernatant.
There is no band in lane 10~12. Use these 3 lanes as negative control.