Paul.xu at 23:31, 4 October 2019

2019-10-04T23:31:18Z

New page

[[File:2616 2615 cloning.JPG|RTENOTITLE]]

[[File:2615 2616 digestion2.JPG|RTENOTITLE]]

http://www.excellgen.net/eln/images/4/4a/2616_2621_digestion.JPG

== expression screening ==

9/27/19

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| Lane
| CLone ID
| Name
| Induction
| Fraction
| Expression
|-
| 1
| 2615
| pCold-MBP-(tev)-GST-(3c)-Ac-LysargiNase 
| 25 oC , 4h
| soluble
| 
|-
| 2
| 2615
| 
| 37 oC 4 h
| soluble 
| Y
|-
| 3
| 2616
| pET28b-hFth-CYFRA 21-1-avitag-6H 
| 25 oC , 4h 
| soluble 
| Y
|-
| 4
| 2616
| 
| 37 oC 4 h 
| soluble 
| Y
|-
| 5
| 2621
| pET29a-NSP4-iFlp 
| 25 oC , 4h 
| soluble 
| 
|-
| 6
| 2621
| 
| 37 oC 4 h 
| soluble 
| 
|-
| 7
| 2615
| pCold-MBP-(tev)-GST-(3c)-Ac-LysargiNase 
| 25 oC , 4h 
| IB
| Y
|-
| 8
| 2615
| 
| 37 oC 4 h 
| IB 
| Y
|-
| 9
| 2616
| pET28b-hFth-CYFRA 21-1-avitag-6H 
| 25 oC , 4h 
| IB 
| Y
|-
| 10
| 2616
| 
| 37 oC 4 h 
| IB 
| Y
|-
| 11
| 2621
| pET29a-NSP4-iFlp 
| 25 oC , 4h 
| IB 
| 
|-
| M
| 
| 1250, 130, 100, 70, 55, 35, 25, 15, 10
| 
| 
| 
|}

[[File:2615 2616 2621 expression screening.jpg|RTENOTITLE]]

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| Lane
| ID
| Name
| Induction
| Sample
| Expression
|-
| 1
| 2555
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC
| 
| 
|-
| 2
| 2556
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 3
| 2557
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 4
| 2558
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 5
| 2559
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 6
| 2560 
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 7
| 2561
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 8
| 1562
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 9
| 2563
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC 
| 
| 
|-
| 10
| 2564
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC, cell lysed 
| 
| 
|-
| 11
| 971
| 
| 5052 o/n 37 oCm 1 mM 3 hours 37 oC, cell lysed 
| 
| 
|-
| M
| 
| 
| 
| 
| 
|-
| 12
| 2621
| pET29a-NSP4-iFlp, 9.76 / 52.8 
| 5052 o/n 37 oCm 1 mM 3 hours 18 oC 
| S 
| 
|-
| 13
| 2615
| pCold-MBP-(tev)-GST-(3c)-Ac-LysargiNase, 5.23 / 110.7 
| 5052 o/n 37 oCm 1 mM 3 hours 18 oC 
| S 
|  
|-
| 14
| 2615
| pCold-MBP-(tev)-GST-(3c)-Ac-LysargiNase 
| 5052 o/n 37 oCm 1 mM 3 hours 18 oC 
| IB
|  
|}

[[File:VLP expression screening 9 28 19.jpg|RTENOTITLE]]

[[File:VLP expression screening 9 28 19 2.jpg|RTENOTITLE]]

== Purification ==

10/4/19

Expression, LB+ 5052, induce at 18 oC o/n, 2 L

10 ml Ni column.

Wash

Elute Low salt Buffer B-->10 ml Q FF

Elute Q FF with 0--> 1.5 M NaCl.

FLPC, 

[[File:2616 Ni.png]]

[[File:2616 Q.png]]http://www.excellgen.net/eln/images/a/af/2616_purification.jpg

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| Lane
| Sample
| 
| 
|-
| 1
| 2616 Ni  frac 3
| 
| 
|-
| 2
| 2616 Ni  frac 4 
| 
| 
|-
| 3
| 2616 Ni  frac 5 
| 
| 
|-
| 4
| 2616 Ni  frac 6 
| 
| 
|-
| 5
| 2616 Ni  frac 12 
| 
| 
|-
| 6
| 2616 Q Frac 13
| 
| 
|-
| 7
| 2616 Q Frac 14 
| 
| 
|-
| 8
| 2616 Q Frac 15 
| 
| 
|-
| M
| Page Ruler Plus
| 
| 
|-
| 9
| 2624 S
| 
| 
|-
| 10
| 2624 secrection
| 
| 
|-
| 11
| ?
| 
| 
|}
[[Category:Cloning]]

2616 - Revision history

Paul.xu at 23:31, 4 October 2019