We have created an extensive part collection in which each piece has a different specific function, however, they all consolidate for the common purpose of creating a flexible and precise nanoplastic detection system.

First, with precise measurements and thorough characterization, we annotated four main parts, which comprise our detection system:

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  BBa_K4380015 → CBD and LCI peptide fusion protein

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  BBa_K4380016 → eGFP and LCI peptide fusion protein

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  BBa_K4380017 → CBD and TA2 peptide fusion protein

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  BBa_K4380018 → eGFP and TA2 peptide fusion protein

Although using known peptides for a detection system already is a huge challenge in itself, we wanted to characterize novel peptides and create huge peptide libraries, which could be used for specific plastic binding in our detection system. Therefore, we decided to do a peptide evolution, specifically designed for plastic-binding peptides.

For this, we created several cell surface display systems, used for the direct evolution of our specific peptides:

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  BBa_K4380019 → EstA protein cell surface display system with mutated LCI peptide

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  BBa_K4380020 → EstA protein cell surface display system with mutated TA2 peptide

These two parts were successfully utilized to create peptide libraries. Although these parts can be used to specifically mutate our peptides, we created a universal system, based on the same protein:

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  BBa_K4380014 → EstA protein cell surface display system

These three cell surface display systems can be successfully amplified with novel error-prone PCR primers, who can be utilized for all of these systems:

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  BBa_K4380006 → epPCR forward primer

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  BBa_K4380007 → epPCR reverse primer

In conclusion, the annotated parts can be successfully used not only for creating a nanoplastic detection system from scratch but also can be utilized for the improvement of proteins, peptides or other parts by direct evolution approaches.