Crm at 16:46, 24 September 2018

2018-09-24T16:46:29Z

New page

[[File:931 digestion.JPG|RTENOTITLE]]

--[[User:Paul.xu|Paul.xu]] ([[User_talk:Paul.xu|talk]]) 15:29, 21 December 2017 (EST)

12/27/2017

938 digestion with Nde1-Xho1, Fastdigest Buffer

[[File:938 939 942 3122 931 1855 digestion.JPG|RTENOTITLE]]

{| border="1" cellspacing="1" cellpadding="1" style="width: 681px;"
|-
| 1
| 2
| 3
| 4
| 5
| 6
| 7
| style="width: 54px;" | 8,9
| style="width: 55px;" | 10,11
| style="width: 71px;" | 13
|-
| 938 PNGase_F
| 939 Tte-ExoIII 
| 942 VVN 
| 3122 CRM197 
| 931, 1, PAK16D ligase  
| 931, 2, PAK16D ligase 
| 1855  yPolyA-Pol 
| style="width: 54px;" | 1540
| style="width: 55px;" | 2046
| style="width: 71px;" | 2046 
|-
| Nde1-Xho1
| Nde1-Xho1 
| Nco1-Xho1 
| Nde1-Xho1 
| Nde1-HindIII 
| HindIII-Xho1
| Nde1-Xho1
| style="width: 54px;" | Nde1-Xho1
| style="width: 55px;" | Nde1-Xho1
| style="width: 71px;" | Nco1-Xho1
|-
| pJet1.2
| pJet1.2 
| pJet1.2 
| pJet1.2 
| pJet1.2 
| pJet1.2 
| pEC2- 
| style="width: 54px;" | pET30b
| style="width: 55px;" | pET28b
| style="width: 71px;" | pET28b 
|-
| 1145 
| 780 
| 716 
| 1798 
| 1174 
| 986 
| 1713
| style="width: 54px;" | there is no Xho1 site
| style="width: 55px;" | 1377
| style="width: 71px;" | ~1377
|-
| 
| 
| 
| 
| 
| 
| 
| style="width: 54px;" | 
| style="width: 55px;" | 
| style="width: 71px;" | 
|}

[[File:938 939 942 3122 931 1855 digestion Cutsmart.JPG|RTENOTITLE]]

== Date:4/5/2018 ==

'''Subclone of 931'''

Vector:1987pT7A3

Insert:

F1:1174 bp

F2:986 bp

[[File:Subclone931.JPG|RTENOTITLE]]

F1:Nde1+HindIII

F2:HindIII+Xho1

=== Date:5/29/2018 ===

'''Subclone of 931'''

'''Vector is 1987pT7A3'''

[[File:IMG 0859.JPG|RTENOTITLE]]

'''Miniprep digestion of 984_931'''

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| #
| 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

[[File:IMG 0871.JPG|RTENOTITLE]]

not good

== Date:7/10/2018 ==

'''931-1 subclone'''

[[File:IMG 1079.JPG|RTENOTITLE]]

'''931 subclone'''

[[File:IMG 1104.JPG|RTENOTITLE]]

'''Miniprep digestion of 931-1'''

[[File:IMG 1091.JPG|RTENOTITLE]]

'''931 miniprep digestion 1'''

[[File:IMG 1127.JPG|RTENOTITLE]]

'''931-1 miniprep digestion'''

Nde1+HindIII

[[File:IMG 1145.JPG|RTENOTITLE]]

'''931subclone'''

[[File:IMG 1147.JPG|RTENOTITLE]]

TE as EB buffer

multiple bands in gel, not good, can not cut out the vector.

== Date:8/17/2018 ==

'''931-1  miniprep'''

vector:pET28b

insert:931-1 in 1727 vector

[[File:IMG 1316.JPG|RTENOTITLE]]

'''931 Subclone'''

vector is 931-1 in pET28b

HindIII+Xho1

[[File:IMG 1318.JPG|RTENOTITLE]]

'''Miniprep digestion of 931'''

[[File:IMG 1373.JPG|RTENOTITLE]]

'''931,#4  digestion'''

{| border="1" cellspacing="1" cellpadding="1" style="width:500px;"
|-
| Lane
| enzyme
| size
|-
| 1
| Nde1+HindIII
| 1174
|-
| 2
| HindIII+Xho1
| 986
|-
| 3
| HindIII+Xho1
| 986
|-
| 4
| Nde1+Xho1
| 2267
|}

[[File:IMG 1377.JPG|RTENOTITLE]]

Junk fragment , from hindIII+Xho1

== Date:9/20/2018 ==

'''931 PCR'''

[[File:IMG 1423.JPG|RTENOTITLE]]

'''931 vector digestion'''

[[File:IMG 1422.JPG|RTENOTITLE]]

'''931 PCR Screening'''

[[File:IMG 1435.JPG]]
[[Category:Cloning]]

931 pET28b-PAK16D ligase - Revision history

Crm at 16:46, 24 September 2018