Crm: Created page with "Spin down 2295 yPolyA. Resuspend 1483 T4 DNA Ligase, homogenize, load onto Ni column. Wash Ni column (bind with 1498 Taq DNA Ligase) with Ni washing buffer, Tris-HCl, Buffer..."

2017-06-14T23:14:47Z

Created page with "Spin down 2295 yPolyA. Resuspend 1483 T4 DNA Ligase, homogenize, load onto Ni column. Wash Ni column (bind with 1498 Taq DNA Ligase) with Ni washing buffer, Tris-HCl, Buffer..."

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Spin down 2295 yPolyA.

Resuspend 1483 T4 DNA Ligase, homogenize, load onto Ni column.

Wash Ni column (bind with 1498 Taq DNA Ligase) with Ni washing buffer, Tris-HCl, Buffer B, dilute and load onto QHP column.

Elute 1498 Taq DNA Ligase with Buffer B 06/14/2017

{| style="width: 500px" cellspacing="1" cellpadding="1" border="1"
|-
| 0.04 
| 0.08 
| 0.95 
| 1.25 
| 1.98 
| (1~5) 
|-
| 2.84
| 4.72 
| 2.16 
| 1.15 
| 0.78 
| (6~10) 
|-
| 0.57 
| 0.41 
| 0.20 
| 0.16 
| 0.05 
| (11~15) 
|}

collect 3~12.

Elute QHP column (load with 1497 ToughTaq)

[[File:128-1497 ToughTaq QHP 6 14 2017.jpg]]

Resuspend 1483 T4 DNA Ligase, homogenize, load onto Ni column.

Run SDS-PAGE gel of 1946 TdT bivine  7.67/45.7 06/14/2017

{| style="width:500px" cellspacing="1" cellpadding="0" border="1" width="500"
|-
|
Number

|
1

|
2

|
3

|
4

|
5

|
6

|
7

|
8

|
9

|
10

|
11

|
12

|-
|
sample

|
18

|
19

|
21

|
23

|
MW

|
24

|
26

|
27

|
28

|
31

|
33

|
34

|-
|
uL

|
5.6

|
4.4

|
1.0

|
1.7

|
10

|
1.3

|
0.7

|
0.9

|
0.9

|
2.5

|
4.8

|
6.7

|-
|
ddH2O

|
10

|
10

|
15

|
15

|
\

|
15

|
16

|
15

|
15

|
10

|
10

|
10

|}

[[File:Protein Expression SDS-PAGE 127-1946 TdT bovine QHP 6 14 2017.jpg]]

Forward sample 18~21 to the next step.

Run SDS-PAGE gel of 1518 T3 Ligase  5.62/41.6     1601 6His-GST-i3C 6.41/50.8     1765 T4 DNA Polymerase 6.19/105.9     1942 phi29 5.72/70.6     06/14/2017

{| style="width:500px" cellspacing="1" cellpadding="0" border="1" width="500"
|-
|
Number

|
1

|
2

|
3

|
4

|
5

|
6

|
7

|
8

|
9

|
10

|
11

|
12

|-
|
sample

|
1518

S

|
1601

S

|
1765

S

|
1942

S

|
MW

|
1518

P

|
1601

P

|
1765

P

|
1942

P

|
1518

U

|
1601

U

|
1765

U

|-
|
uL

|
15

|
15

|
15

|
15

|
10

|
15

|
15

|
15

|
15

|
15

|
15

|
15

|}

S:supernatant

P:sediment

U:uninduced

[[File:Protein Screening 1518 1601 1765 1942 06 14 2017.jpg]]

Only 1518 T3 Ligase and 1942 phi29 can do protein expression.

[[Category:My ELN]]

MS 6 14 2017 - Revision history

Crm: Created page with "Spin down 2295 yPolyA. Resuspend 1483 T4 DNA Ligase, homogenize, load onto Ni column. Wash Ni column (bind with 1498 Taq DNA Ligase) with Ni washing buffer, Tris-HCl, Buffer..."