SPINE - Revision history

Fabian Commichau: /* Studies that made use of SPINE: */

2015-03-09T09:18:12Z

‎Studies that made use of SPINE:

2014-03-31T15:42:54Z

‎Studies that made use of SPINE:

2014-01-09T08:55:42Z

‎A detailed protocol to detect the interaction between RocG and GltC:

2014-01-09T08:53:03Z

‎Studies that made use of SPINE:

2013-12-12T19:09:21Z

‎SPINE for membrane proteins:

2013-12-12T13:50:55Z

‎Studies that made use of SPINE:

2013-12-12T13:50:07Z

‎Studies that made use of SPINE:

2013-07-23T12:42:04Z

‎Studies that made use of SPINE:

2013-07-23T12:38:23Z

‎See the principle

2013-01-16T16:52:21Z

‎A detailed protocol to detect the interaction between RocG and GltC:

@@ Line 38: / Line 38: @@
 =='''Studies that made use of SPINE:'''==
-<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 24375102 24666271 </pubmed>
+<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 24375102 24666271 25711804 </pubmed>
 =='''The use of SPINE in other microbes:'''==
 <pubmed> 21123179 </pubmed>

@@ Line 38: / Line 38: @@
 =='''Studies that made use of SPINE:'''==
-<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 24375102 </pubmed>
+<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 24375102 24666271 </pubmed>
 =='''The use of SPINE in other microbes:'''==
 <pubmed> 21123179 </pubmed>

@@ Line 6: / Line 6: @@
 =='''A detailed protocol to detect the interaction between [[RocG]] and [[GltC]]:''' ==
-litre of a ''B. subtilis'' culture was grown to an OD600 of approx. 1.0 and incubated with 0.6% formaldehyde ( 4% stock solution in PBS, pH 6.5!) for 20 minutes @ 37°C on a shaker.
+litre of a ''B. subtilis'' culture was grown to an OD<sub>600</sub> of approx. 1.0 and incubated with 0.6% formaldehyde ( 4% stock solution in PBS, pH 6.5!) for 20 minutes @ 37°C on a shaker.
 The cells were harvested and washed once in 1 X PBS pH 6.5.
 The pellets can then be stored @ -20 °C.

@@ Line 38: / Line 38: @@
 =='''Studies that made use of SPINE:'''==
-<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 </pubmed>
+<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 22512862 24325460 24375102 </pubmed>
 =='''The use of SPINE in other microbes:'''==
 <pubmed> 21123179 </pubmed>

@@ Line 35: / Line 35: @@
 =='''SPINE for membrane proteins:'''==
-<pubmed> 21472855 </pubmed>
+<pubmed> 21472855 24300168 </pubmed>
 =='''Studies that made use of SPINE:'''==

@@ Line 39: / Line 39: @@
 =='''Studies that made use of SPINE:'''==
 '''Additional references:''' {{PubMed|22512862}}
-<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 23192352 </pubmed>
+<pubmed> 17608797  19193632 20572937 20933603 21622759 21803996 21992469 22001508 22517742 23192352 </pubmed>
 =='''The use of SPINE in other microbes:'''==
 <pubmed> 21123179 </pubmed>

← Older revision		Revision as of 12:38, 23 July 2013
Line 2:		Line 2:


−	==[http://www.iba-lifesciences.com/~~protein_interaction_spine_Technology~~.html '''See the principle''' ]==	+	==[http://www.iba-lifesciences.com/IBA-Applications-Protein-interaction-SPINE-Technology.html '''See the principle''' ]==

	=='''A detailed protocol to detect the interaction between [[RocG]] and [[GltC]]:''' ==		=='''A detailed protocol to detect the interaction between [[RocG]] and [[GltC]]:''' ==

@@ Line 11: / Line 11: @@
 The [[GltC]] protein was expressed carrying a Strep-tag and [[RocG]] expression was induced by arginine ([http://www.ncbi.nlm.nih.gov/sites/entrez/17608797 PubMed]). Expression of the Strep-tagged GltC protein allows to test the functionality of the protein.
 Crude extracts (10-15 ml) were prepared by using a French Press.
-After a centrifugation step for 1 h @ 27.000 g the clarified crude extracts were loaded onto a Streptactin sepharose column (0.5-1 ml matrix) to isolate the cross-linked protein complexes (the detailed procedure for protein purification is described in the IBA manual, http://www.iba-lifesciences.com/protein_interaction_spine_Technology.html).
+After a centrifugation step for 1 h @ 27.000 g the clarified crude extracts were loaded onto a Streptactin sepharose column (0.5-1 ml matrix) to isolate the cross-linked protein complexes (the detailed procedure for protein purification is described in the IBA manual,  http://www.iba-go.com/).
 After the purification of the protein complexes the crosslinks can be resolved by boiling the samples in Laemmli buffer for 10-15 minutes @ 95 °C ([http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Abstract&list_uids=+17994626 PubMed]).
 A 12.5% SDS gel was loaded with the samples and the proteins were then visualized by silver-staining. The interaction partner/s were identified by mass spectroscopy and Western blotting.