Team:UIUC Illinois/Composite Part

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Part Documentation

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• Part: K1681000
• Part Name: PL lac O-1 SCRIBE(kanR)ON
• Part Type: n/a
• Creator: UIUC_Illinois_2015
• Short Description: IPTG-inducible kanamycin resistance SCRIBE cassette
• Long Description: SCRIBE(kanR)ON is a cassette coding for the RNA and proteins needed to restore kanamycin resistance by synthesizing multicopy single-stranded DNA (msDNA) for site-specific recombination. The part consists of the IPTG-inducible promoter PL lac O-1, a retron coding for an untranslated RNA molecule and the open reading frame for reverse transcriptase, a RBS, the open reading frame for beta recombinase, and a terminator. The reverse transcriptase uses the msDNA (msd) region of the RNA molecule as a template and the msd RNA (msr) region of the RNA molecule as a primer to synthesize the msDNA. The beta recombinase inserts the kanamycin resistance segment of the msDNA into the host’s genome during DNA replication as an Okazaki fragment.
• Design Considerations: This part was designed to be used as a positive control for testing SCRIBE cassettes with different promoters and/ or custom DNA segments for site-specific recombination.

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• Part: K1681001
• Part Name: SCRIBE(BsaI)
• Part Type: n/a
• Creator: UIUC_Illinois_2015
• Short Description: SCRIBE cassette with BsaI sites for Golden Gate assembly
• Long Description: SCRIBE(BsaI) is a SCRIBE cassette including two restriction sites of the Type-II restriction enzyme BsaI in the msd region of the retron. This part is compatible with Golden Gate assembly.
• Design Considerations: This part was designed as a modular assembly part for future users to add custom DNA segments into the msd region for site-specific recombination via Golden Gate assembly and to add promoters of their choice via Standard or 3A assembly. See Bba_K1681000 for a more detailed description as to how such custom DNA segments would be inserted into the host’s genome.

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• K1681002
• Part Name: PL lac O-1 SCRIBE(BsaI)
• Part Type: n/a
• Creator: UIUC_Illinois_2015
• Short Description: IPTG-inducible promoter and BsaI SCRIBE cassette
• Long Description: This part is the IPTG-inducible promoter PL lac O-1 immediately upstream of SCRIBE(BsaI), with no scar site in between.
• Design Considerations: This part was designed to be used as a positive control for testing SCRIBE cassettes with different promoters after using Golden Gate assembly to add custom DNA segments into the msd region for site-specific recombination. See Bba_K1681000 for a more detailed description as to how such custom DNA segments would be inserted into the host’s genome.