Part:BBa_K5410014:Design
T7 promoter
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
Here are a few design considerations you need to address during the detailed design of gene editing sequences:
1. Off-target effects: When designing the guide RNA (gRNA) or editing tool, it is critical to minimize off-target binding. This requires careful sequence selection to ensure the gRNA binds only to the intended region, avoiding unintended mutations elsewhere in the genome.
2. PAM sequence selection: The Protospacer Adjacent Motif (PAM) is essential for CRISPR-Cas systems to recognize the target DNA sequence. The choice of PAM sequence affects the target range and efficiency of the editing, so identifying compatible and accessible PAM sites is crucial.
Source
The T7 promoter is a strong, specific promoter recognized by the T7 RNA polymerase, an enzyme encoded by the T7 phage. It drives high levels of transcription in the presence of T7 RNA polymerase, making it commonly used in cloning and protein expression systems. The source of the T7 promoter is the T7 bacteriophage genome, specifically from the region controlling transcription initiation for viral genes.
The source of the HokD terminator would be plasmids or chromosomal regions containing the hok/sok locus, typically found in E. coli and related bacteria. This terminator ensures proper regulation of the hokD gene, which is involved in plasmid stability by selectively inducing cell death in plasmid-free cells.