Part:BBa_K1689010
N-luc-dCas9
Nluc-dCas9 fusion protein
Nluc-dCas9 fusion protein ORF
A catalytically dead Cas9 (dCas9), when coexpressed with a guide RNA, generates a DNA recognition complex which can binds to any targeted gene [1]. And luciferase, widely used as a reporter is split into two fragments, namely Nluc and Cluc [2]. Each protein fragment by itself is inactive, when two fragments are reassembled, the enzymatic activity of the original protein would be reconstituted, providing easily measurable read out. Thus, split luciferase is widely used as a reporter.
2015 Peking iGEM fused Nluc to N terminus of dCas9 (Figure 1). Guided by sgRNA, it binds to target DNA sequence. And together with Cluc-dCas9 (BBa_K1689008):sgRNA complex, our paired dCas9 (PC) reporter system was constructed (Figure 2) which converts invisible sequence information into measurable bioluminescence.
[[File: Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 2451
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 3
Illegal BamHI site found at 4730 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
//awards/part_collection/2015
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