Difference between revisions of "Part:BBa K389004"

Latest revision as of 14:57, 26 October 2010

Luciferase from pGL4.10[luc2]

Structure of Photinus pyralis firefly luciferase. Image taken from: http://en.wikipedia.org/wiki/Luciferase.

A firefly (Photinus pyralis) luciferase from Promega's pGL4.10[luc2] vector. Luciferase is a sensitive reporter gene.

Usage and Biology

The activity of a luciferase is measured in a luminometer - so only use this BioBrick if you have one in your lab.

Firefly luciferase oxidizes its substrate, firefly luciferin, in presence of ATP and oxygen, emitting light ([http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6VSR-4CJB7RR-7&_user=2459438&_coverDate=03%2F31%2F1996&_rdoc=1&_fmt=high&_orig=search&_origin=search&_sort=d&_docanchor=&view=c&_acct=C000057302&_version=1&_urlVersion=0&_userid=2459438&md5=ac60aa60eaad17b2aa1c4b31f954d0a8&searchtype=a Conti et al., 1996]). Because the firefly luciferin does not pass the cell membrane of E. coli under "normal" cultivation conditions the cells have to be lysated before the luciferase activity measurement.

Important parameters

Experiment	Result
Behaviour during cultivation	production is growth dependent degradation in stationary growth phase
Kinetic of luciferin conversion	max. output between 20 - 40 s
Limit of detection (LOD)	162 RLU ~ 0.3 % of BBa_J23103 output
Limit of quantification (LOQ)	306 RLU ~ 0.7 % of BBa_J23103 output

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 64
Illegal NgoMIV site found at 1408
Illegal NgoMIV site found at 1429
Illegal AgeI site found at 1132
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI.rc site found at 1314

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 A firefly (''Photinus pyralis'') luciferase from Promega's pGL4.10[luc2] vector. Luciferase is a sensitive reporter gene.
 ===Usage and Biology===
@@ Line 14: / Line 15: @@
 Firefly luciferase oxidizes its substrate, firefly luciferin, in presence of ATP and oxygen, emitting light ([http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6VSR-4CJB7RR-7&_user=2459438&_coverDate=03%2F31%2F1996&_rdoc=1&_fmt=high&_orig=search&_origin=search&_sort=d&_docanchor=&view=c&_acct=C000057302&_version=1&_urlVersion=0&_userid=2459438&md5=ac60aa60eaad17b2aa1c4b31f954d0a8&searchtype=a Conti ''et al.'', 1996]). Because the firefly luciferin does not pass the cell membrane of ''E. coli'' under "normal" cultivation conditions the cells have to be lysated before the luciferase activity measurement.
+===Important parameters===
+{|cellpadding="10" style="border-collapse: collapse; border-width: 1px; border-style: solid; border-color: #000"
+|-
+!style="border-style: solid; border-width: 1px"| Experiment
+!style="border-style: solid; border-width: 1px"| Result
+|-
+|style="border-style: solid; border-width: 1px"| [[Part:BBa_K389004:Experience#Accumulation of luciferase | Behaviour during cultivation]]
+|style="border-style: solid; border-width: 1px"|
+* production is growth dependent
+* degradation in stationary growth phase
+|-
+|style="border-style: solid; border-width: 1px"| [[Part:BBa_K389004:Experience#Kinetic of luciferin conversion | Kinetic of luciferin conversion]]
+|style="border-style: solid; border-width: 1px"| max. output between 20 - 40 s
+|-
+|style="border-style: solid; border-width: 1px"| [[Part:BBa_K389004:Experience#Sensitivity | Limit of detection (LOD)]]
+|style="border-style: solid; border-width: 1px"| 162 RLU ~ 0.3 % of <partinfo>J23103</partinfo> output
+|-
+|style="border-style: solid; border-width: 1px"| [[Part:BBa_K389004:Experience#Sensitivity | Limit of quantification (LOQ)]]
+|style="border-style: solid; border-width: 1px"| 306 RLU ~ 0.7 % of <partinfo>J23103</partinfo> output
+|}
 <html>