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A 20 bp target sequence and PAM to make guide operator 3
The [http://2016.igem.org/Team:BostonU BostonU 2016] iGEM team synthesized guide operator sequences composed of a 20 base pair target sequence and a PAM sequence. The target sequence was made using a random DNA generator and entered into the [http://crispr.mit.edu/ CRISPR Design Optimization] tool for the human genome (hg19). It was chosen among approximately 1000 other randomly generated target sequences due to its high optimization score ( > 98%) and high expression in an initial screen containing iRFP (figure 1).
The target sequence is expressed by a guide RNA (gRNA) expression vector (BBa_K1875011) from the BostonU Gemini Library. The gRNA binds to dCas9-VPR that targets a paired gRNA operator reporter (BBa_K1875014) to activate a downstream gene of interest (figure 2).
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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