Difference between revisions of "Part:BBa K3739020"
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===Biology=== | ===Biology=== | ||
| − | LCIKR-2 is a mutant of a peptide called LCI from ''Bacillus subtilis'', can sticks to polypropylene strongly | + | LCIKR-2 is a mutant of a peptide called LCI from ''Bacillus subtilis'', can sticks to polypropylene strongly. GFP is green fluorescent protein from jellyfish ''Aequorea Victoria'', which has been widely used as reporter for decades. GFP is fused at C terminal with LCIKR-2 so that the expression of LCIKR-2 will be reported. |
| + | |||
| + | ===Usage=== | ||
| + | Here, we use BBa_3739020 to construct the expression system, which may adhere to polypropylene. | ||
===Characterization=== | ===Characterization=== | ||
====1. Identification==== | ====1. Identification==== | ||
| − | + | In order to verify whether the LCIKR-2-GFP was expressed accurately, ''LCIKR-2-GFP'' (BBa_K3739020) was assembled into the plasmid backbone (Fig. 1A). After being verified by agarose gel electrophoresis, the plasmids have been transformed into ''Vibrio natriegens'' successfully (Fig. 1B). | |
| − | "https://static.igem.org/mediawiki/parts/ | + | "https://static.igem.org/mediawiki/parts/2/2b/T--XMU-China--K3739050.png" |
| − | ::'''Fig. 1.''' Gene circuit involved in protein expression and AGE analysis of string | + | ::'''Fig. 1.''' Gene circuit involved in protein expression and AGE analysis of string (A) Gene circuit was constructed by BBa_K3739020 to express ''LCIKR-2-GFP''. (B) Target bands of ''LCIKR-2-GFP'' (black arrow, 1200 bp). |
| + | |||
| + | ====Proof of the expression==== | ||
| + | After successful construction, the plasmid was transformed into Vibrio natriegens through electroporation. The single positive colony was cultivated in 10 mL LB medium with chloramphenicol (working concentration was 12.5 μg/mL). After ultrasonication broke and centrifugation, the crude protein was released to the supernatant. After being verified by electrophoresed on a sodium dodecyl sulfate (SDS)-10% (wt/vol) polyacrylamide gel (Fig. 2), the protein of LCIKR-2-GFP was successfully expressed. | ||
| + | |||
| + | https://static.igem.org/mediawiki/parts/7/77/T--XMU-China--K3739050P.png | ||
| + | |||
| + | Fig. 2. SDS-PAGE analysis of LCIKR-2-GFP. Target bands of LCIKR-2-GFP (blank arrow, 33.2 kDa). | ||
Latest revision as of 19:47, 21 October 2021
Status: 500
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LCIKR-2 is a mutant of a peptide called LCI from Bacillus subtilis, can sticks to polypropylene strongly. We use Status: 500
Content-type: text/html
Software error:
Not enough arguments for Range::range at /websites/parts.igem.org/cgi/lib/Range.pm line 250, near "range;" Unmatched right curly bracket at /websites/parts.igem.org/cgi/lib/Range.pm line 251, at end of line syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 251, near "}" Can't use global @_ in "my" at /websites/parts.igem.org/cgi/lib/Range.pm line 261, near "= @_" syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 282, near "}" Can't use global @_ in "my" at /websites/parts.igem.org/cgi/lib/Range.pm line 286, near "= @_" Global symbol "$course" requires explicit package name at /websites/parts.igem.org/cgi/lib/Range.pm line 288. syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 314, near "}" Compilation failed in require at /websites/parts.igem.org/cgi/lib/Part.pm line 16. BEGIN failed--compilation aborted at /websites/parts.igem.org/cgi/lib/Part.pm line 16. Compilation failed in require at /websites/parts.igem.org/cgi/partsdb/putout.cgi line 8. BEGIN failed--compilation aborted at /websites/parts.igem.org/cgi/partsdb/putout.cgi line 8.
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to verify its capability by purification the protein.
Biology
LCIKR-2 is a mutant of a peptide called LCI from Bacillus subtilis, can sticks to polypropylene strongly. GFP is green fluorescent protein from jellyfish Aequorea Victoria, which has been widely used as reporter for decades. GFP is fused at C terminal with LCIKR-2 so that the expression of LCIKR-2 will be reported.
Usage
Here, we use BBa_3739020 to construct the expression system, which may adhere to polypropylene.
Characterization
1. Identification
In order to verify whether the LCIKR-2-GFP was expressed accurately, LCIKR-2-GFP (BBa_K3739020) was assembled into the plasmid backbone (Fig. 1A). After being verified by agarose gel electrophoresis, the plasmids have been transformed into Vibrio natriegens successfully (Fig. 1B).
"
"
- Fig. 1. Gene circuit involved in protein expression and AGE analysis of string (A) Gene circuit was constructed by BBa_K3739020 to express LCIKR-2-GFP. (B) Target bands of LCIKR-2-GFP (black arrow, 1200 bp).
Proof of the expression
After successful construction, the plasmid was transformed into Vibrio natriegens through electroporation. The single positive colony was cultivated in 10 mL LB medium with chloramphenicol (working concentration was 12.5 μg/mL). After ultrasonication broke and centrifugation, the crude protein was released to the supernatant. After being verified by electrophoresed on a sodium dodecyl sulfate (SDS)-10% (wt/vol) polyacrylamide gel (Fig. 2), the protein of LCIKR-2-GFP was successfully expressed.
Fig. 2. SDS-PAGE analysis of LCIKR-2-GFP. Target bands of LCIKR-2-GFP (blank arrow, 33.2 kDa).
Sequence and Features
Status: 500
Content-type: text/html
Software error:
Not enough arguments for Range::range at /websites/parts.igem.org/cgi/lib/Range.pm line 250, near "range;" Unmatched right curly bracket at /websites/parts.igem.org/cgi/lib/Range.pm line 251, at end of line syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 251, near "}" Can't use global @_ in "my" at /websites/parts.igem.org/cgi/lib/Range.pm line 261, near "= @_" syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 282, near "}" Can't use global @_ in "my" at /websites/parts.igem.org/cgi/lib/Range.pm line 286, near "= @_" Global symbol "$course" requires explicit package name at /websites/parts.igem.org/cgi/lib/Range.pm line 288. syntax error at /websites/parts.igem.org/cgi/lib/Range.pm line 314, near "}" Compilation failed in require at /websites/parts.igem.org/cgi/lib/Part.pm line 16. BEGIN failed--compilation aborted at /websites/parts.igem.org/cgi/lib/Part.pm line 16. Compilation failed in require at /websites/parts.igem.org/cgi/partsdb/putout.cgi line 8. BEGIN failed--compilation aborted at /websites/parts.igem.org/cgi/partsdb/putout.cgi line 8.
For help, please send mail to this site's webmaster, giving this error message and the time and date of the error.