Pre Lab 3.2 Flashcards
1
Q
Recombinant protein tag
A
6x-10x histidine tag
2
Q
Metal resin that immobilize tag
A
Nickel/ other metals for purification like cobalt
3
Q
Ni-NTA meaning
A
Nickel-nitrilotriacetic acid
(Nitrilo tri acetic)
4
Q
Purpose Ni-NTA
A
Resin containing affinity tag
ni-nta agarose
5
Q
Reagents
A
Binding buffer (0-20 mM imidazole ph 8.0)
Wash buffer (10-20 mM imidazole ph 8.0)
Elution buffer (50-250 mM imidazole ph 8.0)
Final wash buffer (300 mM imidazole ph 8.0)
All have 300 mM NaCl, mM NaH2PO4
6
Q
Imidazole
A
Side chain of histidine
Pentagon with N and NH
C3H4N2
7
Q
Purpose of buffers
A
Binding buffer - associate/prepare for protein
Wash buffer - remove excess and unbound components
Elution buffer - elute/remove bound histidine tagged target proteins
Final wash
